Abstract:
The present invention relates to a method for optimizing the assembled result of sequencing data using a genetic map. In particular, provided in the present invention is a new method for assembling individual sequenced segments, which comprises the step of constructing the genetic map with a genetic marker. Furthermore, also provided in the present invention is a method for assembling the individual sequenced segments into a genome sequence, such as a chromosome sequence.

Description:
FIELD 
       [0001]    The present disclosure relates to fields of genetic engineering technology, genetics and bioinformatics, and more particularly to a method of optimizing an assembly result of sequencing data using a genetic map. Thus, the present disclosure provides a novel method of assembling reads of an individual, comprising a step of constructing a genetic map using genetic markers. In addition, the present disclosure also provides a method of assembling genomic sequencing data into genomic sequence, such as a chromosomal sequence. 
       BACKGROUND 
       [0002]    The Next-Generation DNA sequencing technology is a high-throughput sequencing technique with low cost, and the principle of the Next-Generation DNA sequencing technology is sequencing by synthesis. As an example, the Solexa sequencing method comprises: firstly, randomly breaking DNA double strands using a physical method; secondly, ligating a specific adaptor to an obtained DNA fragments at both ends, in which the specific adaptor comprises an amplifying primer sequence; thirdly, sequencing DNA fragments ligating the specific adaptor. During sequencing, a DNA polymerase synthesizes a complementary strand to a fragment to be tested using an adaptor, and detect a fluorescence signal carried by the newly incorporated bases to obtain a base sequence, then the sequence of the fragment to be tested may be obtained. These obtained sequences are known as reads. A basic process of the Solexa sequencing method may refer to, for example, http:/www.illumina.com. 
         [0003]    To restore a whole sequence status of a genome (for example, assembling reads into a genomic sequence, such as a chromosome sequence), the Next-Generation sequencing method generally takes the way of connecting reads in gradients. Firstly, using an overlapping relationship between reads, the reads are extended as much as possible (i.e. connecting together), to form contigs. Secondly, using a distance relationship between both ends of the reads in a double-ended sequencing, different contigs of the reads comprising two ends are connected together by adding a certain number of N in the middle, and these fragments so formed are known as scaffolds. In each scaffold, a sequential relationship of the contigs about the N-region is already known, and a distance in the DNA sequence thereof is also known. Lastly, an information of these N-regions is restored into ATCG by means of “filling holes”. One method of “filling holes” comprises: finding a double-ended sequencing read with one end falling into a known sequence of the scaffold, and the other end falling into the N-region of the scaffold; calculating all reads falling into the N-region; and partially assembling using the overlapping relationship to obtain the sequence information of the N-region. A general process of sequence connecting may refer to, for example, Li, R. et al. De novo assembly of human genomes with massively parallel short read sequencing.  Genome Res  20, 265-72 (2010). 
         [0004]    Although a known software may be used to connect the sequencing data (i.e. reads) obtained using the Next-Generation sequencing method, a length of reads obtained using the Next-Generation sequencing method are generally shorter (generally only 100 nt). Thus, there is a certain limitation when performing data connecting: it is difficult to connect the reads to form the genomic sequence, such as the chromosome sequence, only relying on assembly software. 
         [0005]    Thus, it is an urgent need to improve the method of assembling the sequencing data (i.e. reads), to further optimize the assembly result of the sequencing data, for example, the reads are connected into the genomic sequence, such as the chromosomal sequence. 
       SUMMARY 
       [0006]    In the present disclosure, unless otherwise stated, technical and scientific terms used herein have commonly understood meanings by those skilled in the art. And, laboratory procedures of genetics, molecular biology and nucleic acid chemistry used herein are all conventional procedures widely used in the related field. At the same time, to better understand the present disclosure, definitions and explanations of relevant terms are provided below. 
         [0007]    As used herein, the term “genetic map” is also known as a linkage map and a chromosomal map, which shows a relative distance (i.e. a genetic distance) between genes or genetic markers, but does not show a physical distance of the genes or the genetic markers on a chromosome. In the genetic map, a position relationship between the genes and the genetic markers is described using the genetic distance, and the genetic distance is calculated using recombination rate. In general, the longer distance between two genes or genetic markers on the same chromosome, the greater probability of a recombination occurrence in the course of meiosis, the smaller probability of a common genetic. Based on a segregation of offspring characters thereof, a recombination rate thereof may be calculated, so as to calculate the genetic distance on the genetic map thereof. In the case of a recombination rate of two genes or genetic markers being 1%, the genetic distance thereof may be define as 1 cm (centimorgan). 
         [0008]    At present, commonly-used genetic markers mainly comprise restriction fragment length polymorphism (RFLP), simple sequence repeats (SSR), sequence-tagged site (STS) and single nucleotide polymorphism (SNP). These genetic markers are all well-known to those killed in the art, which may refer to, for example, Agarwal, M., Shrivastava, N. &amp; Padh, H. Advances in molecular marker techniques and their applications in plant sciences.  Plant cell reports  27, 617-631 (2008). 
         [0009]    As used herein, term “SNP” refers to a polymorphism of DNA sequence caused by a variation of a single nucleotide. SNP is the most common one among biological heritable variations, which accounts at least 90% of all known polymorphisms. SNP site widely exists in a genome of each species. Specifically, in human genome, averagely every 500 to 1,000 base-pairs having an SNP site, and the total number thereof may be estimated up to 3 million or more. 
         [0010]    As used herein, term “reads” refers to sequencing data obtained using various sequencing methods to perform sequencing. For example, a Next-Generation sequencing method, such as 
         [0011]    Solexa sequencing method is an optimized method for providing reads. 
         [0012]    As used herein, term “scaffold” refers to fragments obtained by connecting the reads by means of an overlapping relationship and a physical distance relationship between the reads. 
         [0013]    As used herein, expression “assembling reads into a chromosomal sequence” refers to clustering together the reads derived from an individual, and arranging them in accordance with their order or relative position on a chromosome (optionally, the reads are firstly connected into the scaffolds, and then the steps of clustering and arranging are performed), to obtain a status of the relative position of each fragment on the chromosome, and then to obtain the chromosomal sequence of the individual or a part of the chromosomal sequence thereof. Accordingly, the expression involves a process of clustering and arranging. In the case of the reads completely covering an entire chromosome, an intact chromosomal sequence may be obtained. However, if the reads cannot cover the entire chromosome, then the status of the relative position of these fragments on the chromosome may be obtained, as well as the part of the chromosomal sequence (i.e. there is a part of the chromosomal sequence still unknown, which needs to be determined by further sequencing). 
         [0014]    As used herein, expression “assemble reads (or scaffolds)” refers to arranging each read (or scaffold) in accordance with a relationship of the relative position. 
         [0015]    As used herein, expression “arrange” not only refers to arranging each read (or scaffold) in accordance with a relationship of the relative position, but also to determining a connecting direction of each fragments. 
         [0016]    In the present disclosure, the inventors combine the genetic map with the assembly of the reads, to provide a novel method of assembling the sequencing data (i.e. reads), which optimizes the assembly result of the sequencing data and enables assembling the reads into the genomic sequence, such as the chromosomal sequence. 
         [0017]    The present disclosure is at least partially based on the following principle: if the genetic distance between two genes or genetic markers is very short, such two genes or genetic markers may be then regarded as being linked. Usually, the physical distance of the two linked genes or genetic markers on a sequence is also close, and the two linked genes or genetic markers belong to a same chromosome. Thus, the linkage relationship between the genetic markers in the genetic map may be used to cluster together the reads or scaffolds, comprising a linked marker, in accordance with the chromosome, and a size relationship and a relative position between the genetic markers may be used to orderly connect the reads or the scaffolds into the chromosomal sequence, or part sequence of the chromosome. 
         [0018]    Specifically, in the present disclosure, the inventors exemplarily construct a genetic map using SNP genetic markers. An obtained genetic map comprises a large amount of the SNP markers, and provides a linkage relationship among these SNP markers. Accordingly, based on the linkage relationship among the SNP markers in the genetic map, reads or scaffolds comprising a linked SNP marker may be clustered together in accordance with a chromosome. Further, based on a genetic distance and a relative position between the SNP markers, the reads or the scaffolds belonged to the same chromosome may be orderly arranged, to realize assembling the reads into the chromosomal sequence. 
         [0019]    Thus, in one aspect, the present disclosure provides a method of assembling reads of an individual. The method may comprise: 
         [0020]    constructing a genetic map using genetic markers, in which the genetic map is used to cluster and arrange the reads comprising the genetic markers, to assemble the reads; 
         [0021]    In a preferred embodiment, optionally, prior to clustering and arranging the reads, the reads are connected into scaffolds, and then the genetic map is used to cluster and arrange the scaffolds. Methods well-known in the art may be used to connect the reads into the scaffold, for example, a Soap Denovo assembly software is used. 
         [0022]    In a preferred embodiment, the genetic markers are SNP site markers. 
         [0023]    In a preferred embodiment, the reads derived from a progeny population of the individual are aligned to the scaffolds of the individual, to search and determine the SNP site markers. 
         [0024]    In a preferred embodiment, a SOAP software and a SOAPSnp software are used to search and determine the SNP site markers. 
         [0025]    In a preferred embodiment, a Next-Generation sequencing method, such as a Solexa sequencing method, is used to sequence a genome of the individual, to obtain the reads of the individual. 
         [0026]    In a preferred embodiment, the individual is an animal (such as mammal) or a plant (such as monocotyledon, dicotyledon and the like) 
         [0027]    In another aspect, the present disclosure provides a method of assembling reads of an individual into a chromosomal sequence. The method may comprise: 
         [0028]    1) providing the reads of the individual; 
         [0029]    2) optionally, connecting the reads into scaffolds; 
         [0030]    3) constructing a genetic map using genetic markers; 
         [0031]    4) determining a linkage relationship between the genetic markers using a genetic distance between the genetic markers in the genetic map, to cluster together the reads or the scaffolds comprising the genetic markers in accordance with a chromosome; 
         [0032]    5) arranging the reads or the scaffolds, belonging to a same chromosome, in a sequential order using the genetic distance between the genetic markers in the genetic map, and determining a connecting direction of each fragment, to assemble the reads into the chromosomal sequence. 
         [0033]    In a preferred embodiment, in step 1), a Next-Generation sequencing method, for example a Solexa sequencing method, is used to sequence a genome of the individual, to provide the reads of the individual; 
         [0034]    In a preferred embodiment, in step 2), a SOAP Denovo assembly software is used to connect the reads into the scaffolds. 
         [0035]    In a preferred embodiment, in step 3), the used genetic markers are SNP site markers. 
         [0036]    In a preferred embodiment, in step 3), the reads derived from a progeny population of the individual are aligned to the scaffolds of the individual, to search and determine the SNP site markers. 
         [0037]    In a preferred embodiment, in step 3), a SOAP software and a SOAPSnp software are used to search and determine the SNP site markers. 
         [0038]    In a preferred embodiment, at least three genetic markers are selected from each read or each scaffold for steps 4) and 5). 
         [0039]    The linkage relationship between the genetic markers may be determined based on methods well-known in the art (See, for example, Botstein, D., White, R. L., Skolnick, M. &amp; Davis, R. W. Construction of a genetic linkage map in man using restriction fragment length polymorphisms.  American Journal of Human Genetics  32, 314 (1980)). 
         [0040]    In a preferred embodiment, in step 4), the linkage relationship between the genetic markers is determined by following steps: 
         [0041]    1) calculating a genetic distance between every two of all genetic markers; 
         [0042]    2) setting a threshold value according to a distribution of all genetic distances, for example the threshold value is set as a minimum of confidence interval being 95% or less (99%) of the distribution; 
         [0043]    wherein two genetic markers of which the genetic distance are below the threshold value are regarded as being linked and belonging to the same chromosome. 
         [0044]    In a preferred embodiment, the same number of the genetic markers (such as at least 3) is selected from each read or each scaffold for step 4), and in step 4), the reads or the scaffolds are clustered together in accordance with the chromosome by following steps: 
         [0045]    1) clustering together the reads or the scaffolds comprising linked genetic markers, to form linkage groups; 
         [0046]    optionally, performing steps 2) and 3): 
         [0047]    2) for all reads or all scaffolds which cannot be clustered together to form any linkage groups in step 1), 
         [0048]    calculating a quadratic sum of a genetic distance of the genetic markers in each unclustered fragment and a genetic distance of the genetic markers in each fragment of all linkage groups respectively; 
         [0049]    selecting an unclustered fragment having a minimal quadratic sum and a corresponding fragment which has been clustered into the linkage groups; and 
         [0050]    clustering the unclustered fragment into the linkage groups the corresponding clustered fragment belonged; 
         [0051]    3) repeating step 2), until a total genetic distance of the linkage groups reaching a genetic map total distance of species the individual belonged; in the case of the genetic map total distance of the species being unknown, clustering all scaffolds into the linkage groups. 
         [0052]    The above-described method may realize clustering most (for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or more) or all of the reads or the scaffolds together in accordance with the chromosome. 
         [0053]    In a preferred embodiment, in step 5), an MSTmap software is used to arrange the genetic markers, to determine the sequential order of each scaffold comprising the genetic markers and belonging to the same chromosome. 
         [0054]    In a preferred embodiment, the individual is an animal (such as mammal) or a plant (such as monocotyledon, dicotyledon and the like). 
         [0055]    In another aspect, the present disclosure provides usage of a genetic marker in assembling reads of an individual. 
         [0056]    In a preferred embodiment, the genetic markers are SNP site markers. 
         [0057]    In a preferred embodiment, the reads of the individual are obtained by sequencing a genome of the individual using a Next-Generation sequencing method, such as a Solexa sequencing method 
         [0058]    In a preferred embodiment, the reads of the individual are firstly connected into scaffolds, for example a SOAPDenovo assembly software is used to connect the reads into the scaffolds, and then further assembly is performed using the genetic markers; 
         [0059]    In a preferred embodiment, the genetic markers are used to assemble the reads of the individual into a chromosomal sequence. 
         [0060]    In a preferred embodiment, the individual is an animal (such as mammal) or a plant (such as monocotyledon, dicotyledon and the like). 
         [0061]    General methods of constructing a genetic map using genetic markers, such as SNP, are known to those skilled in the art. (See, for example Shifman, S. et al. A high-resolution single nucleotide polymorphism genetic map of the mouse genome.  PLoS biology  4, e395 (2006) and Groenen, M. A. M. et al. A high-density SNP-based linkage map of the chicken genome reveals sequence features correlated with recombination rate.  Genome research  19, 510 (2009)). In the present disclosure, SNP is taken as an example, which exemplarily provides a method of constructing a genetic map. 
         [0062]    For constructing SNP genetic map, it is usually needed to determine SNP site and calculate genetic distance (i.e. recombination rate) between each SNP site. Accordingly, a progeny population of the target individual, into which the reads are assembled, usually are firstly obtained (for example, the target individual as a parent hybridizes with a reference, and then is subjected to self-breeding, to provide the progeny population), and then the SNP site is determined and the genetic distance between each SNP site is calculated by means of such progeny population (i.e. recombination rate) 
         [0063]    Determination of the SNP Site 
         [0064]    Taken plant as an example, a plurality of individuals in the progeny population of the target individual, into which the reads are assembled, are sequenced. In general, a sequencing depth of each progeny individual is about 2× to 3× (i.e. the total data volume of the reads reaches to about 2-3 times) or more, to basically cover the entire genomic sequence. Thus, respective sequencing data of the plurality of progeny individuals from the target individual may be obtained (i.e. reads). 
         [0065]    Then using, for example, a SOAP software (Li, R. et al. SOAP2: an improved ultrafast tool for short read alignment. Bioinformatics 25, 1966-7 (2009)), the reads of each progeny individual are aligned back into the parent which is obtained by connecting into the scaffolds (i.e. the target individual); and for example, a SOAPSNP software (Li, R. et al. SNP detection for massively parallel whole-genome resequencing.  Genome Research  19, 1124 (2009)) is used to search a SNP site (i.e. a site comprising a difference of a single base between a parent individual and a progeny individual). 
         [0066]    Prior to performing alignment, optionally, the reads of each progeny individual may be filtered, to remove unqualified reads in each individual. The unqualified reads include but not limited to following cases: 
         [0067]    the base number, which a sequencing quality is below a certain threshold value (determined by specific technique and sequencing environment), exceeds 50% of the base number of all reads; 
         [0068]    the base (i.e. N of the reads) number with uncertain sequencing result in the reads exceeds 5% of the base number of all reads; 
         [0069]    an exogenous sequence presented in the reads (an introduced exogenous sequence by experiment, for example, except an adaptor sequence of a sample). 
         [0070]    when performing alignment, a default parameter of a software is generally used, without an allowance of a gap existence, and the number of mismatching is not more than 5 bases. In addition, those reads which can be aligned to a plurality of sites in a genome are generally filtered out. 
         [0071]    Furthermore, the SOAPSNP result is subjected to processing, to search those SNP sites which exist in parent but segregate in progeny. Scaffolds, which these SNP sites locate, and coordinates thereof in the scaffolds are both recorded. The process of searching and determining the SNP site is shown in  FIG. 1 . 
         [0072]    Calculation of a Genetic Distance Between SNP Sites 
         [0073]    According to the information of the SNP site of each progeny individual, a base of the SNP site in the progeny individual derived from a male parent or a female parent (i.e. genotype information) may be determined, which can further determine a distribution of the base of the SNP site in the parent individual among all progeny individuals (See  FIG. 2 ). Thus, a recombination rates between every two SNP sites can be calculated, to obtain a genetic distance between any two SNP sites. The genetic distance is calculated using a mapping function described in Kosambi, D. The estimation of map distances from recombination values.  Annals of Human Genetics  12, 172-175 (1943), in which M represents the genetic distance, r represents the recombination rate, then: 
         [0000]    
       
         
           
             M 
             = 
             
               
                 1 
                 4 
               
                
               
                 ln 
                  
                 
                   ( 
                   
                     
                       1 
                       + 
                       
                         2 
                          
                         
                             
                         
                          
                         r 
                       
                     
                     
                       1 
                       - 
                       
                         2 
                          
                         
                             
                         
                          
                         r 
                       
                     
                   
                   ) 
                 
               
             
           
         
       
       
         
           
             r 
             = 
             
               
                 ( 
                 
                   1 
                   - 
                   
                     same 
                     total 
                   
                 
                 ) 
               
               / 
               2 
             
           
         
       
     
         [0074]    in which, same is the number of which two bases of the SNP site derive from the same parent individual, total is the total number of the individuals. 
         [0075]    According to the above formula, the genetic distance between every two SNP sites may be calculated, which may further construct a SNP genetic map. On this basis, a linkage relationship between every two SNP marker sites may be determined Normally, two SNP sites of which genetic distances are very close are regarded as being linked, and the physical distance thereof in the chromosome is not too far, i.e., such two SNP sites may be basically regarded as belonging to the same chromosome. 
         [0076]    Clustering of the Scaffolds 
         [0077]    On the basis of constructed genetic map, by means of the relative position relationship and the linkage relationship between the genetic markers in the genetic map, the scaffolds of the parent individual (target individual) may be clustered in accordance with a chromosome. An exemplary method of clustering the scaffolds in accordance with the chromosome is provided below. 
         [0078]    In order to simply the complexity of analysis, it may not need to subject all searched SNP sites to clustering. In general, three SNP site markers may be selected from each scaffold: in which two of them locate at two ends of the scaffolds respectively (one locates at a front-end of the scaffold, and the other locates at a back-end of the scaffold), while the third SNP site marker locates in the middle of the scaffold. The genetic distances between the SNP site located in the middle of the scaffold and several SNP sites surrounding are usually not very long, and two SNP site markers located at two ends of the scaffold close to the every end of the scaffold as much as possible, and the genetic distance between these two SNP site markers is greater than zero. 
         [0079]    The genetic distance between every two SNP sites is calculated, the total number of pairwise SNP site markers with equal genetic distance is subjected to statistics, with which a graph is plotted taken the genetic distance as X-coordinate and taken the total number of pairwise SNP site markers as Y-coordinate. Using qqplot (Wilk, M. B. &amp; Gnanadesikan, R. Probability plotting methods for the analysis of data.  Biometrika  55, 1 (1968)) function of R software, it has been found that the distribution of the above plotted graph follows Normal Distribution. An abscissa value of the distribution of which a confidence interval is at least 95% is taken as a threshold value, and two SNP site markers of which the abscissa value is less than the threshold value are regarded as belonging to the same chromosome. 
         [0080]    Thus, if the genetic marker between two SNP site markers, which locate at different scaffolds, being less than the threshold value, then these two scaffolds are regarded as belonging to the same chromosome. Based on this, all scaffolds may be clustered, and those scaffolds clustered together are regarded as a linked group. 
         [0081]    In some cases, there may be some scaffolds which cannot be clustered into any linkage group. In these cases, the scaffolds which cannot be clustered into any linkage group may need to be further clustered into the linkage groups. Accordingly, following method may be used for further clustering: 
         [0082]    1) calculating a quadratic sum of a genetic distance of the genetic markers in each unclustered fragment and a genetic distance of the genetic markers in each fragment of all linkage groups respectively; 
         [0083]    selecting an unclustered fragment having a minimal quadratic sum and a corresponding fragment which has been clustered into the linkage groups; and 
         [0084]    clustering the unclustered fragment into the linkage groups the corresponding clustered fragment belonged; 
         [0085]    2) repeating step 1) until a total genetic distance of the linkage groups reach genetic map total distance of species the individual belonged (if the genetic map total distance of the species being unknown, all scaffolds are clustered into the linkage groups), which may realize clustering the scaffolds, which cannot be clustered into any linkage group, into the linkage group. Thus, all scaffolds or at least most scaffolds (for example, at least 50% of the scaffolds, at least 60% of the scaffolds, at least 70% of the scaffolds, at least 80% of the scaffolds, at least 90% of the scaffolds, at least 95% of the scaffolds, at least 96% of the scaffolds, at least 97% of the scaffolds, at least 98% of the scaffolds, at least 99% of the scaffolds, or more scaffolds) of the parent individual (the target individual) may be clustered. 
         [0086]    Sorting of the Scaffolds 
         [0087]    After clustering the scaffolds, the genetic distance between the genetic markers (for example, the SNP site marker) may be used to sort various scaffolds belonged to the same chromosome. For example, an MSTmap software (Wu, Bhat et al. 2008) may be used to sort the SNP site marker located in the middle of the scaffold. The MSTmap software may be able to sort various scaffolds by constructing a minimum spanning tree, according to a size of the genetic distances between various genetic markers. In general, an actual sequential order of the genetic marker may be obtained by calculating a minimum spanning tree of the graph. Based on this, a relative relationship of various genetic markers, which locate in the middle of the scaffolds, in the linkage group may be obtained, which may further determine the sequential order of various scaffolds belonged to the same chromosome. 
         [0088]    Determination of a Connected Direction of the Scaffolds 
         [0089]    Furthermore, the genetic distance between the genetic markers (such as SNP site marker) may be used to determine a connected direction of various scaffolds. 
         [0090]    For example, after sorting various scaffolds belonged to the same chromosome, a genetic distance between the SNP site markers located at both ends of one scaffold (front-end and back-end) and the SNP site marker located in the middle of previous scaffold, which may determine the connected direction of the one scaffold with the previous scaffold. If a genetic distance between the SNP site marker located at either end of the scaffold and the SNP site marker located in the middle of the previous scaffold is relatively close, then the very end of the one scaffold connects to the previous scaffold, which may determine the connected direction of the one scaffold. Optionally, any other suitable marker combination (for example, SNP site markers located at front-end and middle of scaffold with a pending connected direction, or SNP site markers located at back-end and in middle of scaffold with a pending connected direction, as well as any one of SNP site markers of the previous scaffold) may be used to determine the connected direction of the scaffold 
         [0091]    After clustering and sorting the scaffolds as well as determining the connected direction (for example, according to above-mentioned steps), most scaffolds may be clustered together and then aligned to a chromosome or a certain fragment of a chromosome, so as to assemble reads into a chromosomal sequence.  FIG. 3  exemplarily shows an assembly result of reads derived from a watermelon which is a species with a smaller genome (11 chromosomes) (the used assembly method is similar with the method described in Examples), in which the left side represents a genetic sequential relationship of the genetic marker, the right side represents a position relationship of the scaffold in the chromosome. Such assembly result proves the reliability and effectiveness of the method of the present disclosure, i.e. the method of the present disclosure may be used to effectively assemble the reads of the individual into the chromosomal sequence. 
       Advantageous Effects of the Present Disclosure 
       [0092]    The present disclosure innovatively combines a genetic map with reads together, to provide a novel method of assembling sequencing data (i.e., reads). Comparing with the prior art, the technical solution of the present disclosure has following advantageous effects: 
         [0093]    1) it has been solved as a choke point that reads are unable to be assembled into a genomic sequence (such as chromosomal sequence) using a reads assembly software, which optimizes an assembly result of sequencing data; 
         [0094]    2) it has been realized that reads are assembled into a genomic sequence, such as a chromosomal sequence, which provides a more powerful tool for genomics. 
         [0095]    Reference and examples will be made in details to embodiments of the present disclosure, and it would be appreciated by those skilled in the art that following figures and examples are explanatory, illustrative and used to generally understand the present disclosure, but not construed to limit the present disclosure. According to following detailed descriptions of figures and preferred embodiments, various purposes and advantages of the present disclosure will become apparent to those skilled in the art. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0096]    These and other aspects and advantages of embodiments of the present disclosure will become apparent and more readily appreciated from the following descriptions made with reference to the accompanying drawings, in which: 
           [0097]      FIG. 1  schematically describes a flow chart of searching SNP site using SOAP software and SOAPSnp software. 
           [0098]      FIG. 2  schematically demonstrates a genotype information of the progeny individual, in which, a represents deriving from male parent, b represents deriving from female parent. 
           [0099]      FIG. 3  schematically demonstrates an assembly result of reads, in which the left side represents a genetic sequential relationship of genetic markers, the right side represents a relationship of scaffolds on a chromosome. 
           [0100]      FIG. 4  is a distribution diagram of genetic markers between SNP site markers derived from 9311 rice, in which X-coordinate represents genetic distance; Y-coordinate represents the total number of pairwise SNP site markers. 
           [0101]      FIG. 5  schematically demonstrates a partial assembly result of reads derived from 9311 rice (i.e. a linkage group LG 09), in which, the left side represents a genetic sequential relationship of genetic markers, the right side represents a position relationship of scaffolds on a chromosome. 
       
    
    
     DETAILED DESCRIPTION 
       [0102]    In order to make the purpose, technical solution and advantage of the present disclosure more apparent, a further description will be described in details to the present disclosure. It would be appreciated by those skilled in the art that specific examples described herein are explanatory for the present disclosure, but not be construed to limit the present disclosure. 
       Example 1 
       [0103]    In the present example, 9311 rice was taken as an example, which exemplarily described the method of assembling reads according to the present disclosure. 
         [0104]    Obtaining Scaffolds of 9311 Rice 
         [0105]    The genome of 9311 rice was sequenced using Solexa sequencing platform (illumine company), to provide reads of 9311 rice. Then, using methods well-known in the art, for example Soap Denovo assembly software (http://soap.genomics.org.cn/soapdenovo.html), the reads of 9311 rice was connected into scaffolds, these sequence information of the scaffolds may refer to Yu, Hu et al. 2002. 
         [0106]    Obtaining Progeny Population of 9311 Rice 
         [0107]    The 9311 rice (Yu, J. et al. A draft sequence of the rice genome ( Oryza sativa  L. ssp. indica).  Science  296, 79 (2002)) was subjected to hybridization with pa64 rice (Wei, G et al. A transcriptomic analysis of superhybrid rice LYP9 and its parents.  Proc Natl Acad Sci USA  106, 7695-701 (2009)), to obtain F1 generation, and then the F1 generation self-bred for 16 generations, to obtain a progeny population of 9311 rice. 135 progeny individuals were selected randomly from the progeny population obtained from self-breeding for 16 generations, to subject to an individual sequencing having a sequencing depth of 2× (a data volume of twice genome), to provide reads of the progeny individual. 
         [0108]    Searching and Determining SNP Site 
         [0109]    Taking the scaffolds from the parent 9311 rice as a reference sequence, using SOAP software (Li, R. et al. SOAP2: an improved ultrafast tool for short read alignment.  Bioinformatics  25, 1966-7 (2009)), the reads of the 135 progeny individuals were aligned back to the reference sequence. 
         [0110]    Based on the aligned result obtained using SOAP software, SOAPSnp software (See, for example, http://soap.genomics.org.cn/soapsnp.html or Li, R. et al. SNP detection for massively parallel whole-genome resequencing.  Genome Research.  19, 1124 (2009)) was used to search SNP site, and determine a genotype of each SNP site in progeny individual (i.e. to determine whether a base of SNP site in the progeny individual derived from the 9311 rice or the pa64 rice). 
         [0111]    A statistical result of SNP site from the 9311 rice was shown as Table. 1 
         [0000]    
       
         
               
             
               
               
               
               
               
               
             
           
               
                 TABLE. 1 
               
             
             
               
                   
               
               
                 The statistical result of SNP site from the 9311 rice 
               
             
          
           
               
                   
                   
                   
                   
                   
                 percentage of  
               
               
                   
                   
                   
                 the total 
                 the total 
                 scaffolds 
               
               
                   
                   
                   
                 number of 
                 length of 
                 comprising 
               
               
                   
                   
                 the total 
                 scaffolds 
                 scaffolds 
                 SNP site 
               
               
                   
                   
                 number of 
                 comprising 
                 comprising 
                 marker in the  
               
               
                   
                   
                 SNP site 
                 SNP site 
                 SNP site 
                 whole length 
               
               
                   
                   
                 marker 
                 marker 
                 marker 
                 of genome 
               
               
                   
                   
               
               
                   
                   
                 45516 
                 537 
                 340306986 bp 
                 89.5% 
               
               
                   
                   
               
             
          
         
       
     
         [0112]    As can be seen from the statistical result in Table. 1, the SNP site marker not only had a huge number, but also had a basically uniform distribution in the entire genome. And, these SNP site markers basically covered the entire genome, so as to use in assembling the scaffolds into a genomic sequence (for example a chromosomal sequence) 
         [0113]      FIG. 2  demonstrated a genotype information of partial SNP sites in progeny individuals, in which a represented deriving from a male parent, b represented deriving from a female parent. Based on these genotype information, a distribution of a base of each SNP site in the progeny individuals were determined, to calculate a recombination rate between SNP site markers. 
         [0114]    Clustering and Arranging Scaffolds 
         [0115]    In order to clustering the scaffolds, three SNP site markers were selected out from each scaffold, in which, two of them located at two ends of the scaffolds respectively (one located at a front-end of the scaffold, and the other located at a back-end of the scaffold), while the third SNP site marker located in the middle of the scaffold. The genetic distances between every two of all selected SNP site markers were calculated. The number of the pairwise SNP site markers having the same genetic distance was subjected to statistics, with which a graph was plotted taken the genetic distance as X-coordinate and taken the number of pairwise SNP site markers as Y-coordinate (See  FIG. 4 ). 
         [0116]      FIG. 4  demonstrated a distribution of genetic markers between SNP site markers in 9311 rice. A qqplot function (Wilk, M. B. &amp; Gnanadesikan, R. Probability plotting methods for the analysis of data.  Biometrika  55, 1 (1968)) of R software was used to subject the distribution to a statistical test. The result showed that the distribution of the genetic distance between the SNP site markers basically followed Normal Distribution (R=0.8863972). 
         [0117]    A 99% confidence interval of the distribution was calculated, of which a lower limit was taken as a threshold value, so as to obtain a genetic distance having a threshold value of about 3 cm. Thus, if a genetic distance between two SNP site markers being less than 3 cm, then these two SNP site markers were regarded as linked, and belonged to a same chromosome. Accordingly, the scaffolds of which these two SNP site markers located were also regarded as belonging to a same chromosome. 
         [0118]    According to the above-described threshold value of genetic distance, all scaffolds were clustered. The results showed that, after clustering, 12 linkage groups were obtained (corresponding to the number of chromosome having a haploid in rice. 
         [0119]    Furthermore, those scaffolds which cannot be clustered together to any linkage groups, were clustered by following steps: 
         [0120]    1) calculating a quadratic sum of a genetic distance of SNP site marker in each unclustered scaffold with SNP site marker in various scaffolds of all linkage groups; selecting an unclustered scaffold having a minimal quadratic sum and a corresponding scaffold which has been clustered into the linkage groups; and clustering the unclustered scaffold to the linkage groups which the corresponding clustered scaffold belonged; 2) repeating step 1), until a total genetic distance of all linkage groups reached a genetic map total distance of rice species. 
         [0121]    According to the above steps, there were total 444 scaffolds had been clustered, the total length of the scaffolds was 338,305,001 bp, which accounted for 88.2% of the genome size. And it had been realized that most scaffolds were clustered together in accordance with the chromosome. 
         [0122]    After the clustering steps were completed, an MSTmap soft (Wu, Y, Bhat, P. R., Close, T. J. &amp; Lonardi, S. Efficient and accurate construction of genetic linkage maps from the minimum spanning tree of a graph.  PLoS Genet  4, e1000212 (2008)) was used to sort the clustered scaffolds, to determine the sequential relationship thereof in the linkage groups. Then, a relative genetic distance between the SNP site marker located at both ends of various scaffolds and the SNP site marker located in the middle of the previous scaffold thereof, to determine a connected direction of various scaffolds. By the above-described assembly method, 12 linkage groups (corresponding to 12 chromosomes of the 9311 rice) were obtained, of which the detailed information had been shown in Table. 2. In addition,  FIG. 5  exemplarily demonstrated an arranging situation of scaffolds in one linkage group (linkage group LG 09 of the 9311 rice, which was corresponding to chromosome 9 of the 9311 rice). To be noted, because the length of the chromosomal sequence obtained by assembling was too long,  FIG. 4  only exemplarily demonstrated partial scaffolds of the linkage group LG 09, but not showed all scaffolds. However, those skilled in the art may obtain the chromosomal sequence comprising all scaffolds according to the information in Table. 2. 
         [0000]    
       
         
               
             
               
               
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                 TABLE. 2 
               
             
             
               
                   
               
               
                 A statistical result of sequential order, length and connected direction of  
               
               
                 scaffolds in 12 linkage groups of the 9311 rice. 
               
             
          
           
               
                 name of 
                   
                   
                   
                   
                   
               
               
                 linkage 
                 sequential 
                 name of 
                 length of 
                 connected direction 
                 corresponding 
               
               
                 groups 
                 order 
                 scaffolds 
                 scaffolds 
                 of scaffolds 
                 chromosome 
               
               
                   
               
             
          
           
               
                 LG 01 
                 1 
                 scaffold002365 
                 35,090 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 2 
                 scaffold009522 
                 3,075 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 3 
                 scaffold002666 
                 18,717 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 4 
                 scaffold000417 
                 22,419 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 5 
                 scaffold000165 
                 58,979 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 6 
                 scaffold000001 
                 18,111,789 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 7 
                 scaffold000069 
                 1,088,650 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 8 
                 scaffold002624 
                 19,409 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 9 
                 scaffold000009 
                 10,573,739 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 10 
                 scaffold000190 
                 39,777 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 11 
                 scaffold003391 
                 9,530 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 12 
                 scaffold002717 
                 17,872 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 13 
                 scaffold000226 
                 35,285 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 14 
                 scaffold003570 
                 8,365 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 15 
                 scaffold000216 
                 35,809 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 16 
                 scaffold007201 
                 3,529 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 17 
                 scaffold000020 
                 5,490,501 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 18 
                 scaffold000511 
                 19,178 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 19 
                 scaffold003404 
                 9,545 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 20 
                 scaffold004156 
                 6,919 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 21 
                 scaffold012513 
                 1,908 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 22 
                 scaffold002747 
                 17,080 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 23 
                 scaffold002816 
                 15,709 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 24 
                 scaffold004927 
                 5,479 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 25 
                 scaffold014965 
                 1,297 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 26 
                 scaffold001954 
                 2,990 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 27 
                 scaffold000457 
                 20,981 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 28 
                 scaffold002954 
                 13,632 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 29 
                 scaffold003080 
                 11,955 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 30 
                 scaffold000011 
                 9,076,302 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 31 
                 scaffold012765 
                 2,169 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 32 
                 scaffold002380 
                 33,420 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 33 
                 scaffold003173 
                 11,199 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 34 
                 scaffold002415 
                 29,546 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 35 
                 scaffold000149 
                 92,299 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 36 
                 scaffold000388 
                 23,633 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 37 
                 scaffold000394 
                 23,424 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 38 
                 scaffold005574 
                 4,876 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 39 
                 scaffold006966 
                 3,979 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 40 
                 scaffold002471 
                 25,958 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 41 
                 scaffold000409 
                 22,602 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 42 
                 scaffold002310 
                 44,766 
                 reverse 
                 chromosome01 
               
               
                 LG 01 
                 43 
                 scaffold001419 
                 5,743 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 44 
                 scaffold000433 
                 21,805 
                 forward 
                 chromosome01 
               
               
                 LG 01 
                 45 
                 scaffold000950 
                 10,391 
                 forward 
                 chromosome01 
               
               
                 LG 02 
                 1 
                 scaffold000014 
                 7,042,807 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 2 
                 scaffold000391 
                 23,509 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 3 
                 scaffold000864 
                 11,691 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 4 
                 scaffold000040 
                 2,598,321 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 5 
                 scaffold000996 
                 9,827 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 6 
                 scaffold000254 
                 33,215 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 7 
                 scaffold002980 
                 13,385 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 8 
                 scaffold002644 
                 19,285 
                 reverse 
                 chromosome02 
               
               
                 LG 02 
                 9 
                 scaffold000302 
                 28,827 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 10 
                 scaffold002279 
                 28,540 
                 reverse 
                 chromosome02 
               
               
                 LG 02 
                 11 
                 scaffold003665 
                 8,221 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 12 
                 scaffold000340 
                 26,191 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 13 
                 scaffold002688 
                 17,899 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 14 
                 scaffold000002 
                 17,331,200 
                 reverse 
                 chromosome02 
               
               
                 LG 02 
                 15 
                 scaffold002449 
                 27,340 
                 reverse 
                 chromosome02 
               
               
                 LG 02 
                 16 
                 scaffold001026 
                 9,481 
                 reverse 
                 chromosome02 
               
               
                 LG 02 
                 17 
                 scaffold000356 
                 25,230 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 18 
                 scaffold000303 
                 28,662 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 19 
                 scaffold000246 
                 33,854 
                 reverse 
                 chromosome02 
               
               
                 LG 02 
                 20 
                 scaffold000026 
                 4,123,896 
                 reverse 
                 chromosome02 
               
               
                 LG 02 
                 21 
                 scaffold002785 
                 16,205 
                 forward 
                 chromosome02 
               
               
                 LG 02 
                 22 
                 scaffold002292 
                 51,983 
                 reverse 
                 chromosome02 
               
               
                 LG 02 
                 23 
                 scaffold000022 
                 5,126,128 
                 forward 
                 chromosome02 
               
               
                 LG 03 
                 1 
                 scaffold000349 
                 25,675 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 2 
                 scaffold002418 
                 29,631 
                 reverse 
                 chromosome03 
               
               
                 LG 03 
                 3 
                 scaffold002763 
                 16,852 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 4 
                 scaffold000913 
                 10,988 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 5 
                 scaffold000027 
                 3,804,194 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 6 
                 scaffold003659 
                 8,205 
                 reverse 
                 chromosome03 
               
               
                 LG 03 
                 7 
                 scaffold002569 
                 21,758 
                 reverse 
                 chromosome03 
               
               
                 LG 03 
                 8 
                 scaffold002778 
                 16,613 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 9 
                 scaffold000085 
                 553,483 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 10 
                 scaffold003242 
                 10,493 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 11 
                 scaffold002275 
                 78,376 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 12 
                 scaffold008308 
                 3,400 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 13 
                 scaffold000505 
                 19,501 
                 reverse 
                 chromosome03 
               
               
                 LG 03 
                 14 
                 scaffold000168 
                 54,450 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 15 
                 scaffold002907 
                 13,617 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 16 
                 scaffold003110 
                 11,720 
                 reverse 
                 chromosome03 
               
               
                 LG 03 
                 17 
                 scaffold001914 
                 3,144 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 18 
                 scaffold003157 
                 11,285 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 19 
                 scaffold000013 
                 7,064,451 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 20 
                 scaffold000019 
                 5,919,547 
                 reverse 
                 chromosome03 
               
               
                 LG 03 
                 21 
                 scaffold000375 
                 23,961 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 22 
                 scaffold000281 
                 30,362 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 23 
                 scaffold000123 
                 156,507 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 24 
                 scaffold000380 
                 23,803 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 25 
                 scaffold000091 
                 500,931 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 26 
                 scaffold000003 
                 14,112,554 
                 forward 
                 chromosome03 
               
               
                 LG 03 
                 27 
                 scaffold000015 
                 6,757,605 
                 reverse 
                 chromosome03 
               
               
                 LG 03 
                 28 
                 scaffold000265 
                 32,034 
                 forward 
                 chromosome03 
               
               
                 LG 04 
                 1 
                 scaffold000016 
                 6,434,379 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 2 
                 scaffold001567 
                 4,903 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 3 
                 scaffold000683 
                 14,989 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 4 
                 scaffold001170 
                 7,791 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 5 
                 scaffold003174 
                 10,348 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 6 
                 scaffold000060 
                 1,310,831 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 7 
                 scaffold000626 
                 16,282 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 8 
                 scaffold003510 
                 8,891 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 9 
                 scaffold000111 
                 309,965 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 10 
                 scaffold000099 
                 425,752 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 11 
                 scaffold000108 
                 331,095 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 12 
                 scaffold002741 
                 17,175 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 13 
                 scaffold002377 
                 21,815 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 14 
                 scaffold002376 
                 10,666 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 15 
                 scaffold002728 
                 17,270 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 16 
                 scaffold000081 
                 626,297 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 17 
                 scaffold007442 
                 3,711 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 18 
                 scaffold003666 
                 8,109 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 19 
                 scaffold000224 
                 35,319 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 20 
                 scaffold002796 
                 16,306 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 21 
                 scaffold000166 
                 57,446 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 22 
                 scaffold002927 
                 14,004 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 23 
                 scaffold000031 
                 3,170,253 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 24 
                 scaffold002319 
                 42,545 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 25 
                 scaffold003458 
                 9,082 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 26 
                 scaffold004211 
                 6,688 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 27 
                 scaffold000055 
                 1,556,420 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 28 
                 scaffold002437 
                 27,999 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 29 
                 scaffold002455 
                 26,970 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 30 
                 scaffold002600 
                 20,569 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 31 
                 scaffold002695 
                 18,201 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 32 
                 scaffold002525 
                 23,814 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 33 
                 scaffold000533 
                 18,352 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 34 
                 scaffold000078 
                 811,129 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 35 
                 scaffold000342 
                 26,047 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 36 
                 scaffold002432 
                 27,682 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 37 
                 scaffold002352 
                 36,948 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 38 
                 scaffold002677 
                 18,259 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 39 
                 scaffold000090 
                 513,098 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 40 
                 scaffold002653 
                 18,939 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 41 
                 scaffold004745 
                 5,566 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 42 
                 scaffold003508 
                 8,809 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 43 
                 scaffold000093  
                 488,138 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 44 
                 scaffold002328 
                 40,792 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 45 
                 scaffold002349 
                 37,321 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 46 
                 scaffold000148 
                 98,390 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 47 
                 scaffold000075 
                 880,192 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 48 
                 scaffold002396 
                 31,546 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 49 
                 scaffold002618 
                 20,088 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 50 
                 scaffold000539 
                 18,200 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 51 
                 scaffold000374 
                 24,098 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 52 
                 scaffold000934 
                 10,687 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 53 
                 scaffold000359 
                 25,060 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 54 
                 scaffold000459 
                 20,888 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 55 
                 scaffold002712 
                 17,664 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 56 
                 scaffold002526 
                 24,010 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 57 
                 scaffold000297 
                 29,077 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 58 
                 scaffold000347 
                 25,686 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 59 
                 scaffold000583 
                 17,240 
                 reverse 
                 chromosome04 
               
               
                 LG 04 
                 60 
                 scaffold000096 
                 442,072 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 61 
                 scaffold000104 
                 391,924 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 62 
                 scaffold000005 
                 13,574,865 
                 forward 
                 chromosome04 
               
               
                 LG 04 
                 63 
                 scaffold000321 
                 27,546 
                 reverse 
                 chromosome04 
               
               
                 LG 05 
                 1 
                 scaffold000057 
                 1,418,651 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 2 
                 scaffold000121 
                 160,616 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 3 
                 scaffold000710 
                 14,337 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 4 
                 scaffold000383 
                 23,761 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 5 
                 scaffold000276 
                 30,719 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 6 
                 scaffold000390 
                 23,570 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 7 
                 scaffold000113 
                 294,440 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 8 
                 scaffold002897 
                 14,395 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 9 
                 scaffold002277 
                 70,998 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 10 
                 scaffold000170 
                 53,093 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 11 
                 scaffold000306 
                 28,406 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 12 
                 scaffold000188 
                 40,249 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 13 
                 scaffold000043 
                 2,387,538 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 14 
                 scaffold001062 
                 8,976 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 15 
                 scaffold005163 
                 5,240 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 16 
                 scaffold002429 
                 27,661 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 17 
                 scaffold001020 
                 9,534 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 18 
                 scaffold000053 
                 1,700,887 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 19 
                 scaffold000088 
                 532,389 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 20 
                 scaffold002814 
                 15,978 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 21 
                 scaffold000084 
                 583,342 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 22 
                 scaffold000176 
                 47,342 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 23 
                 scaffold000061 
                 1,287,921 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 24 
                 scaffold000008 
                 11,869,943 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 25 
                 scaffold000161 
                 64,820 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 26 
                 scaffold000307 
                 28,370 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 27 
                 scaffold000411 
                 22,530 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 28 
                 scaffold000076 
                 859,805 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 29 
                 scaffold000130 
                 139,717 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 30 
                 scaffold000156 
                 72,785 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 31 
                 scaffold002372 
                 34,049 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 32 
                 scaffold004187 
                 6,832 
                 reverse 
                 chromosome05 
               
               
                 LG 05 
                 33 
                 scaffold000012 
                 7,625,277 
                 forward 
                 chromosome05 
               
               
                 LG 05 
                 34 
                 scaffold000362 
                 25,032 
                 forward 
                 chromosome05 
               
               
                 LG 06 
                 1 
                 scaffold002411 
                 30,323 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 2 
                 scaffold006178 
                 4,443 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 3 
                 scaffold000225 
                 35,285 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 4 
                 scaffold002387 
                 32,462 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 5 
                 scaffold002400 
                 31,195 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 6 
                 scaffold003313 
                 10,185 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 7 
                 scaffold002298 
                 49,666 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 8 
                 scaffold002314 
                 43,555 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 9 
                 scaffold000360 
                 25,057 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 10 
                 scaffold011106 
                 2,567 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 11 
                 scaffold000036 
                 2,676,551 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 12 
                 scaffold002979 
                 13,093 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 13 
                 scaffold000115 
                 275,107 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 14 
                 scaffold002936 
                 13,816 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 15 
                 scaffold005295 
                 5,101 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 16 
                 scaffold000041 
                 2,491,508 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 17 
                 scaffold000420 
                 22,376 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 18 
                 scaffold003261 
                 10,441 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 19 
                 scaffold007170 
                 3,864 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 20 
                 scaffold002457 
                 27,132 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 21 
                 scaffold004072 
                 6,959 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 22 
                 scaffold002334 
                 39,311 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 23 
                 scaffold002417 
                 29,224 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 24 
                 scaffold000287 
                 29,960 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 25 
                 scaffold001643 
                 4,450 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 26 
                 scaffold005976 
                 4,180 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 27 
                 scaffold004978 
                 5,475 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 28 
                 scaffold002843 
                 15,265 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 29 
                 scaffold000379 
                 23,821 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 30 
                 scaffold000044 
                 2,330,599 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 31 
                 scaffold000047 
                 2,243,037 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 32 
                 scaffold000032 
                 2,952,239 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 33 
                 scaffold000466 
                 20,558 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 34 
                 scaffold001363 
                 6,114 
                 reverse 
                 chromosome06 
               
               
                 LG 06 
                 35 
                 scaffold000018 
                 5,962,590 
                 forward 
                 chromosome06 
               
               
                 LG 06 
                 36 
                 scaffold000796 
                 12,476 
                 forward 
                 chromosome06 
               
               
                 LG 07 
                 1 
                 scaffold000007 
                 12,232,608 
                 forward 
                 chromosome07 
               
               
                 LG 07 
                 2 
                 scaffold000100 
                 422,751 
                 forward 
                 chromosome07 
               
               
                 LG 07 
                 3 
                 scaffold000056 
                 1,491,444 
                 forward 
                 chromosome07 
               
               
                 LG 07 
                 4 
                 scaffold000038 
                 2,632,557 
                 reverse 
                 chromosome07 
               
               
                 LG 07 
                 5 
                 scaffold000017 
                 6,341,531 
                 forward 
                 chromosome07 
               
               
                 LG 07 
                 6 
                 scaffold000132 
                 133,160 
                 reverse 
                 chromosome07 
               
               
                 LG 08 
                 1 
                 scaffold000077 
                 831,649 
                 forward 
                 chromosome08 
               
               
                 LG 08 
                 2 
                 scaffold000039 
                 2,622,754 
                 forward 
                 chromosome08 
               
               
                 LG 08 
                 3 
                 scaffold000052 
                 1,939,947 
                 reverse 
                 chromosome08 
               
               
                 LG 08 
                 4 
                 scaffold000042 
                 2,466,211 
                 forward 
                 chromosome08 
               
               
                 LG 08 
                 5 
                 scaffold002531 
                 23,148 
                 forward 
                 chromosome08 
               
               
                 LG 08 
                 6 
                 scaffold000033 
                 2,885,658 
                 forward 
                 chromosome08 
               
               
                 LG 08 
                 7 
                 scaffold000079 
                 679,419 
                 reverse 
                 chromosome08 
               
               
                 LG 08 
                 8 
                 scaffold001056 
                 9,104 
                 forward 
                 chromosome08 
               
               
                 LG 08 
                 9 
                 scaffold000006 
                 12,426,518 
                 forward 
                 chromosome08 
               
               
                 LG 08 
                 10 
                 scaffold000035 
                 2,789,649 
                 reverse 
                 chromosome08 
               
               
                 LG 09 
                 1 
                 scaffold002847 
                 15,370 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 2 
                 scaffold000184 
                 42,473 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 3 
                 scaffold000885 
                 11,343 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 4 
                 scaffold000124 
                 155,546 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 5 
                 scaffold002311 
                 44,466 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 6 
                 scaffold000107 
                 342,017 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 7 
                 scaffold006214 
                 4,362 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 8 
                 scaffold000183 
                 42,811 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 9 
                 scaffold000263 
                 32,117 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 10 
                 scaffold005816 
                 3,889 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 11 
                 scaffold002812 
                 16,028 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 12 
                 scaffold000253 
                 33,220 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 13 
                 scaffold000070 
                 1,021,785 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 14 
                 scaffold002406 
                 30,529 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 15 
                 scaffold000211 
                 36,077 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 16 
                 scaffold004084 
                 7,044 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 17 
                 scaffold002494 
                 25,660 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 18 
                 scaffold003540 
                 8,725 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 19 
                 scaffold000222 
                 35,399 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 20 
                 scaffold000850 
                 11,820 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 21 
                 scaffold003302 
                 10,138 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 22 
                 scaffold000337 
                 26,355 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 23 
                 scaffold002271 
                 88,941 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 24 
                 scaffold000063 
                 1,240,123 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 25 
                 scaffold002641 
                 19,323 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 26 
                 scaffold002528 
                 23,662 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 27 
                 scaffold002300 
                 49,469 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 28 
                 scaffold000645 
                 15,731 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 29 
                 scaffold002915 
                 14,144 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 30 
                 scaffold000110 
                 310,809 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 31 
                 scaffold002478 
                 25,752 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 32 
                 scaffold000072 
                 940,878 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 33 
                 scaffold000059 
                 1,319,559 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 34 
                 scaffold002312 
                 43,866 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 35 
                 scaffold000509 
                 19,380 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 36 
                 scaffold002866 
                 15,039 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 37 
                 scaffold003034 
                 12,576 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 38 
                 scaffold002362 
                 36,159 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 39 
                 scaffold002382 
                 33,767 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 40 
                 scaffold001327 
                 6,323 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 41 
                 scaffold002586 
                 20,319 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 42 
                 scaffold000357 
                 25,196 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 43 
                 scaffold002422 
                 28,035 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 44 
                 scaffold003130 
                 11,504 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 45 
                 scaffold002551 
                 22,471 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 46 
                 scaffold002295 
                 51,718 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 47 
                 scaffold000106 
                 376,199 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 48 
                 scaffold000566 
                 17,626 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 49 
                 scaffold002459 
                 26,858 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 50 
                 scaffold002906 
                 13,978 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 51 
                 scaffold000071 
                 973,574 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 52 
                 scaffold000255 
                 33,044 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 53 
                 scaffold002767 
                 16,418 
                 forward 
                 chromosome09 
               
               
                 LG 09 
                 54 
                 scaffold000004 
                 13,648,413 
                 reverse 
                 chromosome09 
               
               
                 LG 09 
                 55 
                 scaffold003102 
                 11,854 
                 reverse 
                 chromosome09 
               
               
                 LG 10 
                 1 
                 scaffold000717 
                 14,199 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 2 
                 scaffold000010 
                 9,226,363 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 3 
                 scaffold002705 
                 17,879 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 4 
                 scaffold002758 
                 16,811 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 5 
                 scaffold000028 
                 3,656,306 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 6 
                 scaffold001106 
                 8,506 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 7 
                 scaffold000339 
                 26,216 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 8 
                 scaffold000080 
                 672,175 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 9 
                 scaffold000145 
                 102,966 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 10 
                 scaffold002395 
                 31,863 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 11 
                 scaffold004664 
                 5,863 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 12 
                 scaffold003373 
                 9,680 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 13 
                 scaffold000049 
                 2,054,425 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 14 
                 scaffold000058 
                 1,347,837 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 15 
                 scaffold000102 
                 400,512 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 16 
                 scaffold003073 
                 12,190 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 17 
                 scaffold000452 
                 21,217 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 18 
                 scaffold002835 
                 15,590 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 19 
                 scaffold002981 
                 13,038 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 20 
                 scaffold003576 
                 8,539 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 21 
                 scaffold003450 
                 9,210 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 22 
                 scaffold002817 
                 15,617 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 23 
                 scaffold002324 
                 41,841 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 24 
                 scaffold003147 
                 10,991 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 25 
                 scaffold003582 
                 8,574 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 26 
                 scaffold000491 
                 19,946 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 27 
                 scaffold002648 
                 19,119 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 28 
                 scaffold000363 
                 24,778 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 29 
                 scaffold003542 
                 8,354 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 30 
                 scaffold002583 
                 21,076 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 31 
                 scaffold002398 
                 31,519 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 32 
                 scaffold003199 
                 10,621 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 33 
                 scaffold002689 
                 18,331 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 34 
                 scaffold000144 
                 107,923 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 35 
                 scaffold002608 
                 20,302 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 36 
                 scaffold000298 
                 29,061 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 37 
                 scaffold004965 
                 5,412 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 38 
                 scaffold002392 
                 32,130 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 39 
                 scaffold002651 
                 19,089 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 40 
                 scaffold000249 
                 33,577 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 41 
                 scaffold000261 
                 32,352 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 42 
                 scaffold000098 
                 436,095 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 43 
                 scaffold014653 
                 1,471 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 44 
                 scaffold007570 
                 3,601 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 45 
                 scaffold002480 
                 26,032 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 46 
                 scaffold000159 
                 70,207 
                 reverse 
                 chromosome10 
               
               
                 LG 10 
                 47 
                 scaffold000037 
                 2,649,063 
                 forward 
                 chromosome10 
               
               
                 LG 10 
                 48 
                 scaffold000352 
                 25,549 
                 forward 
                 chromosome10 
               
               
                 LG 11 
                 1 
                 scaffold000024 
                 4,558,429 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 2 
                 scaffold000064 
                 1,206,036 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 3 
                 scaffold000177 
                 47,109 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 4 
                 scaffold000082 
                 611,242 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 5 
                 scaffold000101 
                 419,278 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 6 
                 scaffold002369 
                 33,986 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 7 
                 scaffold000087 
                 539,582 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 8 
                 scaffold000089 
                 524,755 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 9 
                 scaffold000147 
                 99,912 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 10 
                 scaffold000095 
                 462,442 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 11 
                 scaffold000455 
                 21,057 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 12 
                 scaffold000023  
                 4,580,783 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 13 
                 scaffold000074  
                 905,087 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 14 
                 scaffold000065  
                 1,195,813 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 15 
                 scaffold003053 
                 12,118 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 16 
                 scaffold002804 
                 15,900 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 17 
                 scaffold002479 
                 25,567 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 18 
                 scaffold004907 
                 5,549 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 19 
                 scaffold002374 
                 34,063 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 20 
                 scaffold000030  
                 3,198,014 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 21 
                 scaffold000437 
                 21,566 
                 reverse 
                 chromosome11 
               
               
                 LG 11 
                 22 
                 scaffold000051  
                 1,959,494 
                 forward 
                 chromosome11 
               
               
                 LG 11 
                 23 
                 scaffold000610 
                 16,727 
                 forward 
                 chromosome11 
               
               
                 LG 12 
                 1 
                 scaffold000135 
                 125,195 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 2 
                 scaffold000092  
                 490,349 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 3 
                 scaffold000086  
                 549,244 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 4 
                 scaffold002268 
                 122,910 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 5 
                 scaffold002304 
                 47,478 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 6 
                 scaffold002278 
                 68,340 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 7 
                 scaffold000021 
                 5,247,386 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 8 
                 scaffold000229 
                 35,107 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 9 
                 scaffold002353 
                 36,841 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 10 
                 scaffold002895 
                 14,478 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 11 
                 scaffold002430 
                 28,447 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 12 
                 scaffold002956 
                 13,651 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 13 
                 scaffold000046  
                 2,288,301 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 14 
                 scaffold000274 
                 30,957 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 15 
                 scaffold002559 
                 22,143 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 16 
                 scaffold003569 
                 8,623 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 17 
                 scaffold000062  
                 1,240,444 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 18 
                 scaffold000218 
                 35,631 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 19 
                 scaffold000197 
                 37,784 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 20 
                 scaffold000670 
                 15,190 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 21 
                 scaffold002307 
                 46,441 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 22 
                 scaffold002787 
                 15,725 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 23 
                 scaffold002572 
                 21,261 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 24 
                 scaffold000678  
                 15,037 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 25 
                 scaffold000169  
                 53,110 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 26 
                 scaffold000120  
                 166,455 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 27 
                 scaffold000127  
                 147,478 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 28 
                 scaffold002486 
                 25,542 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 29 
                 scaffold000122  
                 159,240 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 30 
                 scaffold003007  
                 12,920 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 31 
                 scaffold002928  
                 14,029 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 32 
                 scaffold002930  
                 14,039 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 33 
                 scaffold000054 
                 1,669,303 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 34 
                 scaffold002383  
                 33,364 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 35 
                 scaffold000116  
                 260,792 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 36 
                 scaffold000327  
                 27,154 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 37 
                 scaffold002296  
                 50,534 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 38 
                 scaffold003085  
                 11,754 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 39 
                 scaffold002359  
                 36,344 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 40 
                 scaffold002851  
                 14,984 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 41 
                 scaffold001243  
                 7,074 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 42 
                 scaffold000240  
                 34,369 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 43 
                 scaffold002614  
                 20,172 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 44 
                 scaffold002680  
                 18,217 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 45 
                 scaffold002879  
                 14,774 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 46 
                 scaffold002370  
                 34,604 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 47 
                 scaffold002339  
                 38,759 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 48 
                 scaffold000126  
                 148,970 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 49 
                 scaffold000343  
                 25,930 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 50 
                 scaffold002485  
                 25,639 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 51 
                 scaffold002589  
                 21,049 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 52 
                 scaffold002623  
                 19,905 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 53 
                 scaffold000097  
                 436,197 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 54 
                 scaffold003636  
                 7,754 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 55 
                 scaffold000251 
                 33,310 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 56 
                 scaffold002424  
                 28,152 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 57 
                 scaffold000322  
                 27,531 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 58 
                 scaffold002818  
                 15,491 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 59 
                 scaffold004368  
                 6,406 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 60 
                 scaffold002342  
                 38,432 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 61 
                 scaffold003369  
                 9,718 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 62 
                 scaffold004674  
                 5,794 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 63 
                 scaffold002274  
                 78,498 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 64 
                 scaffold000131  
                 139,459 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 65 
                 scaffold000066 
                 1,188,804 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 66 
                 scaffold000048 
                 2,107,733 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 67 
                 scaffold002378  
                 33,507 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 68 
                 scaffold002815  
                 15,332 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 69 
                 scaffold002654  
                 17,840 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 70 
                 scaffold002281  
                 64,592 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 71 
                 scaffold003126  
                 11,466 
                 forward 
                 chromosome12 
               
               
                 LG 12 
                 72 
                 scaffold000025 
                 4,281,268 
                 reverse 
                 chromosome12 
               
               
                 LG 12 
                 73 
                 scaffold000105  
                 390,192 
                 reverse 
                 chromosome12 
               
               
                   
               
             
          
         
       
     
         [0123]    As can be seen from the above result, the method of the present example using a genetic map comprising SNP site marker, broke through the choke point that the Next-Generation sequencing technique-based assembly software cannot connect reads into chromosomal sequence, and successfully realized connecting the reads of the 9311 rice genome into the chromosomal sequence, which provided a more powerful tool for the genomics. 
         [0124]    In addition, the above-describe method was also used to assemble the reads of individual derived from watermelon which is a species with a smaller genome (11 chromosomes). The assembly result of such individual reads was shown in  FIG. 3 , in which the left side represented the genetic sequential relationship of the genetic markers, the right side represented the position relationship of the scaffolds in the chromosome. This assembly result further proved the reliable and effectiveness of the method of the present disclosure, i.e., the method of the present disclosure may be used to effectively assemble the individual reads into the chromosomal sequence. 
         [0125]    Although specific embodiments of the present disclosure have been described in details, the above embodiments cannot be construed to limit the present disclosure. And, it would be appreciated by those skilled in the art that various modification and changes can be made in the embodiments according to all teachings which has been already disclosed, which are all within the scope of the present disclosure. The full scope of the present disclosure is given by the claims and any equivalents thereof. 
         [0126]    In the present text, additional details of publications and other materials for illustrating the present disclosure or providing implement of the present disclosure are all incorporated herein by reference, and following references are provided for convenience. 
         [0127]    1. Kosambi, D. (1944). “The estimation of map distances from recombination values.”  Ann. Eugen.  12: 172-175. 
         [0128]    2. Li, R., Y. Li, et al. (2009). “SNP detection for massively parallel whole-genome resequencing.”  Genome Research  19(6): 1124. 
         [0129]    3. Li, R., Y. Li, et al. (2008). “SOAP: short oligonucleotide alignment program.”  Bioinformatics  24(5): 713. 
         [0130]    4. Li, R., H. Zhu, et al. (2010). “De novo assembly of human genomes with massively parallel short read sequencing.”  Genome Research  20(2): 265. 
         [0131]    5. Wu, Y., P. R. Bhat, et al. (2008). “Efficient and Accurate Construction of Genetic Linkage Maps from the Minimum Spanning Tree of a Graph.”  PLoS Genet  4(10): e1000212. 
         [0132]    6. Yu, J., S. Hu, et al. (2002).“A draft sequence of the rice genome ( Oryza sativa  L. ssp. indica).”  Science  296 (5565): 79. 
         [0133]    7. Li, R. et al. De novo assembly of human genomes with massively parallel short read sequencing.  Genome Res  20, 265-72 (2010). 
         [0134]    8. Agarwal, M., Shrivastava, N. &amp; Padh, H. Advances in molecular marker techniques and their applications in plant sciences.  Plant cell reports  27, 617-631 (2008). 
         [0135]    9. Botstein, D., White, R.L., Skolnick, M. &amp; Davis, R. W. Construction of a genetic linkage map in man using restriction fragment length polymorphisms.  American Journal of Human Genetics  32, 314 (1980). 
         [0136]    10. Shifman, S. et al. A high-resolution single nucleotide polymorphism genetic map of the mouse genome.  PLoS biology  4, e395 (2006). 
         [0137]    11. Groenen, M. A. M. et al. A high-density SNP-based linkage map of the chicken genome reveals sequence features correlated with recombination rate.  Genome research  19, 510 (2009). 
         [0138]    12. Li, R. et al. SOAP2: an improved ultrafast tool for short read alignment.  Bioinformatics  25, 1966-7 (2009). 
         [0139]    13. Li, R. et al. SNP detection for massively parallel whole-genome resequencing.  Genome Research  19, 1124 (2009). 
         [0140]    14. Kosambi, D. The estimation of map distances from recombination values.  Annals of Human Genetics  12, 172-175 (1943). 
         [0141]    15. Wilk, M. B. &amp; Gnanadesikan, R. Probability plotting methods for the analysis for the analysis of data.  Biometrika  55, 1 (1968). 
         [0142]    16. Wu, Y., Bhat, P. R., Close, T. J. &amp; Lonardi, S. Efficient and accurate construction of genetic linkage maps from the minimum spanning tree of a graph.  PLoS Genet  4, el000212 (2008). 
         [0143]    17. Wei, G et al. A transcriptomic analysis of superhybrid rice LYP9 and its parents.  Proc Natl Acad Sci USA  106, 7695-701 (2009).