BACKGROUND
the modern cultivated soybean , which contains high protein and oil content, is an important crop worldwide.
soybean was domesticated from its wild progenitor, glycine soja sieb.
& zucc.
~ <dig>  years ago  <cit> .
although the cultivated and wild soybeans show little reproductive isolation and have very similar genomes in both size and content  <cit> , they exhibit substantial morphological difference .
the pre-domesticated wild soybean accessions  have weedy prostrate growth habits and small black seeds, and the domesticated landraces produce smaller plants with less vegetative growth and often are slightly prostrate.
in contrast, the elite cultivars developed by modern breeding practices have erect and compact stem architecture with reduced branching, high harvest indices, and high seed yield.
the emergence of cultivated crops from their wild progenitors was achieved primarily by artificial selection for a wide range of desirable traits to meet human needs  <cit> .
although domestication traits were often controlled by a relatively small number of genes, including major quantitative trait loci  and/or mendelian loci, selection for such traits would have resulted in a progressive reduction of genetic diversity throughout the genome  <cit> .
genetic diversity was further reduced following domestication by modern breeding practices  <cit> .
the genetic bottlenecks associated with the domestication and genetic improvement of soybean had been illustrated by analysis of  <dig> fragments from  <dig> genes  <cit> .
to date, several agronomically important genes including the dt <dig> locus controlling soybean stem growth habit and e genes  controlling flowering time have been cloned by homology-based or map-based approaches  <cit> .
nevertheless, little is known about how genetic diversity across the whole genome of soybean was shaped by domestication.
the availability of the soybean genome sequence  <cit>  and high throughput sequencing technologies provides an unprecedented opportunity to track the evolutionary history of domesticated soybean, and to dissect the genetic bases for soybean domestication and varietal diversification.
recently, for example,  <dig> soybean accessions, representing wild and cultivated gene pools, had been re-sequenced and analyzed  <cit> .
this study shed light on the nature and extent of genetic differentiation between wild and cultivated soybean species.
nevertheless, no information about landraces – the bridge between wild soybean  and elite cultivars  was provided.
investigations of the loss and recovery of genetic diversity in the course of soybean domestication and breeding would provide guidelines and strategies for utilization of landraces and/or wild accessions for soybean enhancement.
moreover, comparative genomics analyses among wild, landrace, and elite soybeans would identify genes under selection.
the knowledge obtained from these analyses will facilitate the introgression of beneficial alleles from wild soybean and landraces to elite cultivars.
in this study, we re-sequenced  <dig> diverse soybean accessions, which represent three distinct gene pools: the pre-domesticated annual wild progenitor species , domesticated local landraces , and modern elite cultivars .
to achieve a more comprehensive analysis, we integrated these re-sequencing data with the re-sequencing data previously generated from  <dig> wild and  <dig> cultivated soybean genomes  <cit> .
our study not only elucidated the trends of molecular diversity, but also identified distinct footprints in the soybean genomes associated with artificial selection during soybean domestication and elite cultivar development.
RESULTS
high quality sequence data was generated for  <dig> diverse soybean accessions
we used  <dig> diverse soybean accessions in this study: eight wild soybeans, eight landraces, and nine modern elite cultivars.
to maximally represent the genetic diversity and wide geographic distribution, this panel of accessions was selected based on intensive molecular and phenotypic characterization, which reflect the major operational taxonomic units  of soybeans in china  <cit>  .
using the genome-wide re-sequencing approach, a total of  <dig>  billion high-quality paired-end reads  were generated , covering  <dig> % of the genome sequences .
to overcome potential ambiguity caused by sample size and low-pass sequencing in detecting snps  <cit> , we downloaded the  <dig> soybean re-sequencing data through the ncbi short read archive .
after calibrating the snp calling quality by all the  <dig> accessions  and discarding singletons and most doubletons according to rigorous filtering criteria  <cit> , we identified  <dig> , <dig> high quality snps in our sequenced accessions , which was slightly lower than that discovered previously in the  <dig> soybean accessions .
among these,  <dig> %  snps were newly reported here.
additionally, we identified  <dig>  small  insertion/deletions , which provide useful markers for mapping genes, and  <dig>  large deletions , with a mean length of  <dig>  bp.
we validated  <dig> snps from ten randomly selected genes using the sanger method, and the accuracy of snp calling reached  <dig> %, suggesting that potential miscalling of snps in this study was minimal.
bayesian clustering revealed introgression of the wild into the cultivated soybeans
phylogenetic relationships of the  <dig> accessions and williams  <dig>  <cit>  were established using another legume model, medicago truncatula <cit>  as an out-group.
the cultivated and the wild soybeans were separated into two groups , suggesting that the domestication event promoted the genetic differentiation within the subgenera soja.
within the cultivated accessions, the lines l <dig>  l <dig>  l <dig> and l <dig> were separated from the other cultivated accessions.
the bayesian clustering approach revealed different degrees of introgression between the cultivated and the wild groups .
it is particularly interesting that the four landraces  with mosaic pattern at k =  <dig> were found to have at least one of the wild traits, such as small seed size, dark seed-coat color, and seed-coat bloom .
in contrast, two wild accessions  showing admixture carrying one of correspopnding typical cultivated phenotypes .
a recent study revealed that the oryza sativa indica, a cultivated rice subspecies, was developed from crosses between the other cultivated rice subspecies, o. sativa japonica and its wild progenitor o. rufipogon <cit>  suggesting that introgression between the wild and cultivated species and re-selection for desirable agronomic traits may be a common process for crop domestication.
further re-sequencing of larger populations of representative wild and cultivated soybeans such as core collections would allow full elucidation of such evolutionary events occurred during soybean domestication.
within the cultivated soybean group, the landraces were not separated from elite cultivars distinctly .
instead, individuals from the same geographical region tended to cluster together, which reflected isolation by distance during evolution and/or parallel selections in similar ecological habitats accompanied by gene flow.
genome diversity was more impacted by domestication than by genetic improvement
number of snps as well as nucleotide diversity substantially decreased throughout the domestication process from the wild to the cultivated soybeans, which was consistent with previous studies  <cit> .
our data revealed that  <dig> , <dig> snps in wild soybean were not polymorphic in the landraces .
of these snps,  <dig> %  were located in the cds regions of genic sequences and  <dig> %  were non-synonymous sites.
in addition, we observed a reduction of 31% and 26% of genetic diversity from the wild soybeans to landraces, as measured by θπ and θw respectively  <cit>  .
these observations contrasted with a previous study, which reported a reduction of nucleotide diversity from g. soja to landraces at 34% and 51%, measured by θπ and θw, respectively  <cit> .
different samples and different sets of genes were investigated in these two studies, which might explain the different levels of reduction of genetic diversity detected in the two studies.
it is hypothesized that modern plant breeding reduces genetic diversity in elite cultivars, consequently jeopardizing future crop improvement  <cit> .
although this conception appears to be true for most crop species, our data showed limited effects of breeding on reduction of genetic diversity.
we found that the elite gene pool harbored a high proportion of the genetic diversity  presented in the landraces , contrasting with a previous study by hyten et al.
<cit> , which demonstrated that the elite cultivars retained 78%  and 72%  of the diversity present in the landraces.
this difference may indeed reflect the relative levels of genetic diversity of the two sets of elite soybean cultivars investigated in both studies.
the number of fixed snps from landraces to elite cultivars  was only half  of the number of fixed snps during domestication .
similar patterns were observed when only one gene component, such as intron, cds, or utr, was analyzed .
together, these observations indicated that the impact of intensive selection by modern soybean breeding on reduction of genetic diversity was less severe than that of selection by the domestication process, suggesting that the wild soybean gene pool was the major reservoir that retained genes/alleles lost during domestication and modern breeding practice.
we would like to point out that this interpretation would be largely affected by the genetic base of ancestral landraces that were used for the development of elite cultivars investigated in this study.
nevertheless, similar observations were also observed in maize.
a recent study by hufford et al.
demonstrated a remarkably weak genome-wide genetic bottleneck by mordern maize breeding  <cit> .
decrease in the haplotype diversity during domestication
the extent of linkage disequilibrium can be interpreted as a measurement of haplotype diversity in a population.
we observed a drastic increase in linkage disequilibrium  across the whole genome from wild to landraces and elite cultivars  pointing to a severe loss of haplotype diversity.
this observation reflects the genetic bottleneck during domestication, which reduced the genetic diversity throughout the genome by eliminating some recombinant lineages.
it is likely that the lower level of outcrossing rate of the cultivated soybean relative to the wild soybean  <cit>  contributed to an increase in ld in the former.
by contrast, the ld pattern of the landraces differed only slightly from modern elite cultivars .
as a result, the resolution of genome-wide association mapping for panels of landraces or elite cultivars was much lower than that for the wild soybeans.
we also observed a large variation in extent of ld among different chromosomes , suggesting that molecular markers designed for genotyping strategies should be specific to genomic regions in association mapping analyses.
for example, relatively low density of markers is needed for the regions with relatively extensive ld.
footprints of domestication in the soybean genome
the loss of genetic diversity during domestication and genetic improvement is likely due to the fixation and sweep of alleles caused by population bottlenecks or artificial selection.
we scanned a combined dataset of  <dig> accessions to identify genome-wide signatures of artificial selection following a bottom-up genetic approach  <cit> .
to detect the reduction of genetic diversity caused by domestication, we employed a sliding window strategy to estimate θπ <cit>  and tajima’s d  <cit> .
the regions that showed significantly lower θπ in landrace relative to the wild group  and significantly lower tajima’s d  in landraces relative to the wild group were considered as putative domestication-related regions.
this approach has been used to study domestication event in silkworms  <cit>  and rice  <cit> .
the genome scan revealed that only  <dig> % of the whole genome , comprising  <dig> regions distributed on individual chromosomes , appeared to have been affected by selection during domestication.
the length of these regions ranged from  <dig> kbp to  <dig> kbp and the polymorphism levels of these regions relative to the whole genome were relatively low .
a total of  <dig> genes were located in the regions with footprints of artificial selection, accounting for  <dig> % of the  <dig>  predicted genes in the cultivated soybean genome  <cit> .
it was reported that some qtls controlling mesdotication-related traits located in syntenic regions among different species  <cit> .
we found that some candidate genes related with soybean domestication detected in this study had homologs, which were also affected by artificial selection in other crops, such as rice and sunflower.
for example, glyma03g <dig> , which is probably involved in the carbohydrate metabolism pathway, was found to be an orthologous gene of grain incomplete filling  <dig> , a domestication gene identified in rice  <cit> .
gif <dig> encodes a cell-wall invertase that regulates sugar levels for cell division and growth during grain development, resulting in higher seed weight – an important trait for rice domestication.
in addition, we found a strong selection signal for glyma03g <dig> , an orthologous gene of terminal flower  <dig> , which experienced selective sweeps in the domestication of sunflower  <cit> .
as the closest paralogous gene of glyma03g <dig>  in soybean, gmtfl <dig>  was identified to control the agronomically important trait indeterminacy , which is associated with soybean domestication and varietal differentiation  <cit> .
nucleotide diversity analysis of  <dig> wild and  <dig> cultivated soybeans detected five snps in the wild population, but none of them were found in the cultivated population, suggesting that glyma03g <dig>  had experienced artificial selection  <cit> .
footprints of intensive breeding in the soybean genome
population branch statistics  is an effective method to detect signatures of recent natural selection  <cit> .
taking wild soybeans as a control in the pbs approach, we found that  <dig> regions were associated with significant signs  of artificial selection by the modern breeding practice .
these regions spanned a total of  <dig>  kbp in length, corresponding to  <dig> % of the whole genome .
of these  <dig> regions,  <dig> were found to harbor a total of  <dig>  genes showing signatures of selection, which account for  <dig> % of all the genes located in these  <dig> regions  <cit> .
no genes were annotated in the remaining  <dig> regions.
the black seed-coat progressively changed to various colors during domestication, with positive selection for yellow in the following improvement.
multiple alleles at the i locus were found to be associated with an unusual cluster of five chalcone synthase genes  that controlled the distribution of seed-coat color by inhibiting coloration over the entire seed coat  <cit> .
in this study, three  of these five candidate chs genes showed strong selection signals.
the evolution of flowering time was crucial for developing cultivars adapted to a wider geographical regions  <cit> .
we found that two genes related to flowering time, gmcry1a  and glyma10g <dig> , exhibited selection signals.
gmcry1a was a major regulator of photoperiodic flowering in soybean and had an important role in determining latitudinal distribution of soybean  <cit>  while glyma10g <dig>  was a homologous gene of constans , which was found to encod a key protein involved in photoperiod sensing in arabidopsis <cit> .
in total,  <dig> % of the annotated genes were impacted by artificial selection for agricultural traits.
polymorphism levels in the detected selection regions were relatively low compared to that of the whole genome .
the percentage of candidate genes impacted by artificial selection was similar to that was estimated in maize   <cit> .
however, this was slightly lower than that reported  by lam et al.
<cit>  probably due to the sampling effects and different analytical methods employed.
only two regions located on gm <dig> and gm <dig> showed selection signatures for both domestication and subsequent modern breeding practice.
the selected genes appeared to be distributed in clusters in certain genomic regions , similar to the distribution pattern of domestication-related qtls defined by qtl mapping  <cit> .
the domestication and improvement related genes were clustered into  <dig> gene families by orthomcl  <cit> .
of these  <dig> genes,  <dig> were shared by both processes.
using the kegg   <cit>  database, potential functions of the selected genes were predicted.
we found that the selected genes were significantly  involved in lipid metabolism, transcription factors, snares , solute carrier family, and transport and catabolism .
growing demand for vegetable oil is a paramount objective of soybean domestication and genetic improvement, which has focused selection toward cultivars with high accumulation of lipids  <cit> .
a high frequency of selected genes involved in lipid metabolism was also observed during both processes .
this indicates that continuous artificial selection had occurred in the pursuit of preferred-quality soybean seed.
these preliminary data would allow us to prioritize further analyses with an emphasis on understanding of the biological functions of selected genes.
similar to described in maize  <cit> , transcription factors were enriched in the candidate genes with selection signaturs, suggesting that these regulatory genes had been the major target of selection.
of the  <dig> domestication-related genes identified in any plant species to date,  <cit> ,  <dig> were transcription factor genes  <cit> .
these genes were responsible for major morphology differentiation between cultivated crops and their progenitors, such as branch  and glume architecture  in maize  <cit> , seed size  and style length  in tomato  <cit> , seed color  genes in wheat  <cit> , six-rowed spike  in barley  <cit> , seed shattering   <cit>  in rice and in cereal including sorghum, rice and maize   <cit> , fruit opening and seed dispersal  in brassicaceae  <cit> .
a recent study accounted well for this observation which observed that the regulatory genes with stronger regulatory action on the other genes are the targets of selection within the complex regulatory networks inferred from a simulation study using a matrix model  <cit> .
discovering genes with an integrated qtl mapping and re-sequencing approach
although genomic regions and genes, most likely affected by artificial selection, had been identified, the functions and phenotypes of these genes remained elusive  <cit> .
to validate footprints of selection during domestication and genetic improvement, we compared the genomic regions with previously mapped qtls, which were identified from interspecific populations and intraspecific populations developed by crossing landrace and cultivar, respectively .
a total of  <dig> candidate domestication regions including  <dig> genes were covered by the mapped domestication qtls or their adjacent regions  <cit> .
important agronomic traits included yield, plant height, lodging, maturity time, seed weight, seed hardness, seed-coat color, and flower color.
and a total of  <dig> candidate improvement regions including  <dig> genes were covered by improvement qtls or their adjacent regions  <cit> .
in addition, the integration of selection regions identified using population genetic analysis method with qtls region identified using an bi-parents populations may be a useful approach to narrow down the broad qtls  <cit> .
we conducted a linkage mapping study in an interspecific f2: <dig> population consisting of two of the parents included in our survey and searched for qtl for seed size, one of the most prevailing domestication phenotypes .
among the detected qtl we observed one at linkage map of lg f , which accounted for  <dig> % of seed size variation.
the genomic distance between the two flanking markers satt <dig> and satt <dig> was  <dig>  mb .
the selection signals were further identified in eight internal regions  using  <dig> kbp sliding windows in the qtl .
within the narrowed regions,  <dig> genes were potentially responsible for seed size variation.
four regions  were identified as footprints of intensive breeding in this qtl region and a seed size qtl was also discovered nearby using an intraspecific cultivated soybean population  <cit> , indicating that artificial selection occurs continuously in or near the qtl in the pursuit of higher production.
we further identified selection signals within another qtl on gm <dig>  which is responsible for the typical soybean domestication trait, seed blooming   <cit>  .
in  <dig>  mb of this qtl region, three nearby candidate domestication regions consist of  <dig> kbp dna were identified .
this approach offers potential application for cloning candidate genes underlying the domestication traits of soybean as well as other crops.
CONCLUSIONS
soybean has undergone a series of selections over time, natural or artificial, intentional or unintentional, leading to the decrease in genetic diversity from the wild progenitor to landraces and from landraces to the modern elite cultivars.
we reported that whole genome re-sequencing analysis enhanced our understanding of genetic diversity in wild and cultivated soybeans, and unraveled the processes how this important legume species was domesticated.
in present study, the strength of genetic bottlenecks caused by domestication and modern breeding were demonstrated.
the continuing reduction of genetic diversity in the cultivated soybean has become a bottleneck for improvement of soybean cultivars.
we currently have unprecedented opportunities to exploit genetic diversity in the wild soybean and landraces for sustainable enhancement of soybeans.
a set of candidate genes/regions were identified, significantly impacted by selection, for constructing preferred traits underlying soybean domestication and genetic improvement.
comparison of candidate domestication and crop improvement-related genes with previous qtl mapping results, as well as their homologs, provides information on potential function of genes under artificial selection.
in particular, we found genes related to seed-coat color, growth habit, flowering time and seed size, which had been confirmed as continuously changing from wild soybeans to landraces and then elite cultivars.
further analysis is required to identify how variation in these candidate genes affect phenotypes using qtl mapping e.g.
in maize  <cit> , association mapping e.g.
in barley  <cit> , gene expression assays e.g.
in sunflower  <cit> , and gene-knock-out methods  <cit> .
our findings, however, promote development of more efficient approaches to identify the genes underlying domestication-related traits.
this study also contribut <dig> to construct a large-scale soybean haplotype map and discover important trait related genes using genome-wide association studies.
our understanding of the nature of genetic diversity in wild and cultivated soybeans, and the impact of domestication and breeding on genome diversity, will aid future breeding of elite cultivars to improve soybean production and meet the increasing worldwide demands for feed, vegetable oil, soyfood and biofuels.

 the modern cultivated soybean, which contains high protein and oil content, is an important crop worldwide. soybean was domesticated from its wild progenitor, glycine soja sieb. & zucc. ~  years ago. although the cultivated and wild soybeans show little reproductive isolation and have very similar genomes in both size and content, they exhibit substantial morphological difference. the pre-domesticated wild soybean accessions have weedy prostrate growth habits and small black seeds, and the domesticated landraces produce smaller plants with less vegetative growth and often are slightly prostrate. in contrast, the elite cultivars developed by modern breeding practices have erect and compact stem architecture with reduced branching, high harvest indices, and high seed yield. the emergence of cultivated crops from their wild progenitors was achieved primarily by artificial selection for a wide range of desirable traits to meet human needs. although domestication traits were often controlled by a relatively small number of genes, including major quantitative trait loci and/or mendelian loci, selection for such traits would have resulted in a progressive reduction of genetic diversity throughout the genome. genetic diversity was further reduced following domestication by modern breeding practices. the genetic bottlenecks associated with the domestication and genetic improvement of soybean had been illustrated by analysis of fragments from genes. to date, several agronomically important genes including the dt locus controlling soybean stem growth habit and e genes controlling flowering time have been cloned by homology-based or map-based approaches. nevertheless, little is known about how genetic diversity across the whole genome of soybean was shaped by domestication. the availability of the soybean genome sequence and high throughput sequencing technologies provides an unprecedented opportunity to track the evolutionary history of domesticated soybean, and to dissect the genetic bases for soybean domestication and varietal diversification. recently, for example, soybean accessions, representing wild and cultivated gene pools, had been re-sequenced and analyzed. this study shed light on the nature and extent of genetic differentiation between wild and cultivated soybean species. nevertheless, no information about landraces – the bridge between wild soybean and elite cultivars was provided. investigations of the loss and recovery of genetic diversity in the course of soybean domestication and breeding would provide guidelines and strategies for utilization of landraces and/or wild accessions for soybean enhancement. moreover, comparative genomics analyses among wild, landrace, and elite soybeans would identify genes under selection. the knowledge obtained from these analyses will facilitate the introgression of beneficial alleles from wild soybean and landraces to elite cultivars. in this study, we re-sequenced diverse soybean accessions, which represent three distinct gene pools: the pre-domesticated annual wild progenitor species, domesticated local landraces, and modern elite cultivars. to achieve a more comprehensive analysis, we integrated these re-sequencing data with the re-sequencing data previously generated from wild and cultivated soybean genomes. our study not only elucidated the trends of molecular diversity, but also identified distinct footprints in the soybean genomes associated with artificial selection during soybean domestication and elite cultivar development.  high quality sequence data was generated for diverse soybean accessions we used diverse soybean accessions in this study: eight wild soybeans, eight landraces, and nine modern elite cultivars. to maximally represent the genetic diversity and wide geographic distribution, this panel of accessions was selected based on intensive molecular and phenotypic characterization, which reflect the major operational taxonomic units of soybeans in china. using the genome-wide re-sequencing approach, a total of billion high-quality paired-end reads were generated, covering % of the genome sequences. to overcome potential ambiguity caused by sample size and low-pass sequencing in detecting snps, we downloaded the soybean re-sequencing data through the ncbi short read archive. after calibrating the snp calling quality by all the accessions and discarding singletons and most doubletons according to rigorous filtering criteria, we identified, high quality snps in our sequenced accessions, which was slightly lower than that discovered previously in the soybean accessions. among these, % snps were newly reported here. additionally, we identified small insertion/deletions, which provide useful markers for mapping genes, and large deletions, with a mean length of  bp. we validated snps from ten randomly selected genes using the sanger method, and the accuracy of snp calling reached %, suggesting that potential miscalling of snps in this study was minimal. bayesian clustering revealed introgression of the wild into the cultivated soybeans phylogenetic relationships of the accessions and williams were established using another legume model, medicago truncatula as an out-group. the cultivated and the wild soybeans were separated into two groups, suggesting that the domestication event promoted the genetic differentiation within the subgenera soja. within the cultivated accessions, the lines l l l and l were separated from the other cultivated accessions. the bayesian clustering approach revealed different degrees of introgression between the cultivated and the wild groups. it is particularly interesting that the four landraces with mosaic pattern at k =  were found to have at least one of the wild traits, such as small seed size, dark seed-coat color, and seed-coat bloom. in contrast, two wild accessions showing admixture carrying one of correspopnding typical cultivated phenotypes. a recent study revealed that the oryza sativa indica, a cultivated rice subspecies, was developed from crosses between the other cultivated rice subspecies, o. sativa japonica and its wild progenitor o. rufipogon suggesting that introgression between the wild and cultivated species and re-selection for desirable agronomic traits may be a common process for crop domestication. further re-sequencing of larger populations of representative wild and cultivated soybeans such as core collections would allow full elucidation of such evolutionary events occurred during soybean domestication. within the cultivated soybean group, the landraces were not separated from elite cultivars distinctly. instead, individuals from the same geographical region tended to cluster together, which reflected isolation by distance during evolution and/or parallel selections in similar ecological habitats accompanied by gene flow. genome diversity was more impacted by domestication than by genetic improvement number of snps as well as nucleotide diversity substantially decreased throughout the domestication process from the wild to the cultivated soybeans, which was consistent with previous studies. our data revealed that, snps in wild soybean were not polymorphic in the landraces. of these snps, % were located in the cds regions of genic sequences and % were non-synonymous sites. in addition, we observed a reduction of 31% and 26% of genetic diversity from the wild soybeans to landraces, as measured by θπ and θw respectively. these observations contrasted with a previous study, which reported a reduction of nucleotide diversity from g. soja to landraces at 34% and 51%, measured by θπ and θw, respectively. different samples and different sets of genes were investigated in these two studies, which might explain the different levels of reduction of genetic diversity detected in the two studies. it is hypothesized that modern plant breeding reduces genetic diversity in elite cultivars, consequently jeopardizing future crop improvement. although this conception appears to be true for most crop species, our data showed limited effects of breeding on reduction of genetic diversity. we found that the elite gene pool harbored a high proportion of the genetic diversity presented in the landraces, contrasting with a previous study by hyten et al., which demonstrated that the elite cultivars retained 78% and 72% of the diversity present in the landraces. this difference may indeed reflect the relative levels of genetic diversity of the two sets of elite soybean cultivars investigated in both studies. the number of fixed snps from landraces to elite cultivars was only half of the number of fixed snps during domestication. similar patterns were observed when only one gene component, such as intron, cds, or utr, was analyzed. together, these observations indicated that the impact of intensive selection by modern soybean breeding on reduction of genetic diversity was less severe than that of selection by the domestication process, suggesting that the wild soybean gene pool was the major reservoir that retained genes/alleles lost during domestication and modern breeding practice. we would like to point out that this interpretation would be largely affected by the genetic base of ancestral landraces that were used for the development of elite cultivars investigated in this study. nevertheless, similar observations were also observed in maize. a recent study by hufford et al. demonstrated a remarkably weak genome-wide genetic bottleneck by mordern maize breeding. decrease in the haplotype diversity during domestication the extent of linkage disequilibrium can be interpreted as a measurement of haplotype diversity in a population. we observed a drastic increase in linkage disequilibrium across the whole genome from wild to landraces and elite cultivars pointing to a severe loss of haplotype diversity. this observation reflects the genetic bottleneck during domestication, which reduced the genetic diversity throughout the genome by eliminating some recombinant lineages. it is likely that the lower level of outcrossing rate of the cultivated soybean relative to the wild soybean contributed to an increase in ld in the former. by contrast, the ld pattern of the landraces differed only slightly from modern elite cultivars. as a result, the resolution of genome-wide association mapping for panels of landraces or elite cultivars was much lower than that for the wild soybeans. we also observed a large variation in extent of ld among different chromosomes, suggesting that molecular markers designed for genotyping strategies should be specific to genomic regions in association mapping analyses. for example, relatively low density of markers is needed for the regions with relatively extensive ld. footprints of domestication in the soybean genome the loss of genetic diversity during domestication and genetic improvement is likely due to the fixation and sweep of alleles caused by population bottlenecks or artificial selection. we scanned a combined dataset of accessions to identify genome-wide signatures of artificial selection following a bottom-up genetic approach. to detect the reduction of genetic diversity caused by domestication, we employed a sliding window strategy to estimate θπ and tajima’s d. the regions that showed significantly lower θπ in landrace relative to the wild group and significantly lower tajima’s d in landraces relative to the wild group were considered as putative domestication-related regions. this approach has been used to study domestication event in silkworms and rice. the genome scan revealed that only % of the whole genome, comprising regions distributed on individual chromosomes, appeared to have been affected by selection during domestication. the length of these regions ranged from kbp to kbp and the polymorphism levels of these regions relative to the whole genome were relatively low. a total of genes were located in the regions with footprints of artificial selection, accounting for % of the predicted genes in the cultivated soybean genome. it was reported that some qtls controlling mesdotication-related traits located in syntenic regions among different species. we found that some candidate genes related with soybean domestication detected in this study had homologs, which were also affected by artificial selection in other crops, such as rice and sunflower. for example, glyma03g, which is probably involved in the carbohydrate metabolism pathway, was found to be an orthologous gene of grain incomplete filling, a domestication gene identified in rice. gif encodes a cell-wall invertase that regulates sugar levels for cell division and growth during grain development, resulting in higher seed weight – an important trait for rice domestication. in addition, we found a strong selection signal for glyma03g, an orthologous gene of terminal flower, which experienced selective sweeps in the domestication of sunflower. as the closest paralogous gene of glyma03g in soybean, gmtfl was identified to control the agronomically important trait indeterminacy, which is associated with soybean domestication and varietal differentiation. nucleotide diversity analysis of wild and cultivated soybeans detected five snps in the wild population, but none of them were found in the cultivated population, suggesting that glyma03g had experienced artificial selection. footprints of intensive breeding in the soybean genome population branch statistics is an effective method to detect signatures of recent natural selection. taking wild soybeans as a control in the pbs approach, we found that regions were associated with significant signs of artificial selection by the modern breeding practice. these regions spanned a total of kbp in length, corresponding to % of the whole genome. of these regions, were found to harbor a total of genes showing signatures of selection, which account for % of all the genes located in these regions. no genes were annotated in the remaining regions. the black seed-coat progressively changed to various colors during domestication, with positive selection for yellow in the following improvement. multiple alleles at the i locus were found to be associated with an unusual cluster of five chalcone synthase genes that controlled the distribution of seed-coat color by inhibiting coloration over the entire seed coat. in this study, three of these five candidate chs genes showed strong selection signals. the evolution of flowering time was crucial for developing cultivars adapted to a wider geographical regions. we found that two genes related to flowering time, gmcry1a and glyma10g, exhibited selection signals. gmcry1a was a major regulator of photoperiodic flowering in soybean and had an important role in determining latitudinal distribution of soybean while glyma10g was a homologous gene of constans, which was found to encod a key protein involved in photoperiod sensing in arabidopsis. in total, % of the annotated genes were impacted by artificial selection for agricultural traits. polymorphism levels in the detected selection regions were relatively low compared to that of the whole genome. the percentage of candidate genes impacted by artificial selection was similar to that was estimated in maize. however, this was slightly lower than that reported by lam et al. probably due to the sampling effects and different analytical methods employed. only two regions located on gm and gm showed selection signatures for both domestication and subsequent modern breeding practice. the selected genes appeared to be distributed in clusters in certain genomic regions, similar to the distribution pattern of domestication-related qtls defined by qtl mapping. the domestication and improvement related genes were clustered into gene families by orthomcl. of these genes, were shared by both processes. using the kegg database, potential functions of the selected genes were predicted. we found that the selected genes were significantly involved in lipid metabolism, transcription factors, snares, solute carrier family, and transport and catabolism. growing demand for vegetable oil is a paramount objective of soybean domestication and genetic improvement, which has focused selection toward cultivars with high accumulation of lipids. a high frequency of selected genes involved in lipid metabolism was also observed during both processes. this indicates that continuous artificial selection had occurred in the pursuit of preferred-quality soybean seed. these preliminary data would allow us to prioritize further analyses with an emphasis on understanding of the biological functions of selected genes. similar to described in maize, transcription factors were enriched in the candidate genes with selection signaturs, suggesting that these regulatory genes had been the major target of selection. of the domestication-related genes identified in any plant species to date,, were transcription factor genes. these genes were responsible for major morphology differentiation between cultivated crops and their progenitors, such as branch and glume architecture in maize, seed size and style length in tomato, seed color genes in wheat, six-rowed spike in barley, seed shattering in rice and in cereal including sorghum, rice and maize, fruit opening and seed dispersal in brassicaceae. a recent study accounted well for this observation which observed that the regulatory genes with stronger regulatory action on the other genes are the targets of selection within the complex regulatory networks inferred from a simulation study using a matrix model. discovering genes with an integrated qtl mapping and re-sequencing approach although genomic regions and genes, most likely affected by artificial selection, had been identified, the functions and phenotypes of these genes remained elusive. to validate footprints of selection during domestication and genetic improvement, we compared the genomic regions with previously mapped qtls, which were identified from interspecific populations and intraspecific populations developed by crossing landrace and cultivar, respectively. a total of candidate domestication regions including genes were covered by the mapped domestication qtls or their adjacent regions. important agronomic traits included yield, plant height, lodging, maturity time, seed weight, seed hardness, seed-coat color, and flower color. and a total of candidate improvement regions including genes were covered by improvement qtls or their adjacent regions. in addition, the integration of selection regions identified using population genetic analysis method with qtls region identified using an bi-parents populations may be a useful approach to narrow down the broad qtls. we conducted a linkage mapping study in an interspecific f2: population consisting of two of the parents included in our survey and searched for qtl for seed size, one of the most prevailing domestication phenotypes. among the detected qtl we observed one at linkage map of lg f, which accounted for % of seed size variation. the genomic distance between the two flanking markers satt and satt was  mb. the selection signals were further identified in eight internal regions using kbp sliding windows in the qtl. within the narrowed regions, genes were potentially responsible for seed size variation. four regions were identified as footprints of intensive breeding in this qtl region and a seed size qtl was also discovered nearby using an intraspecific cultivated soybean population, indicating that artificial selection occurs continuously in or near the qtl in the pursuit of higher production. we further identified selection signals within another qtl on gm which is responsible for the typical soybean domestication trait, seed blooming. in  mb of this qtl region, three nearby candidate domestication regions consist of kbp dna were identified. this approach offers potential application for cloning candidate genes underlying the domestication traits of soybean as well as other crops.  soybean has undergone a series of selections over time, natural or artificial, intentional or unintentional, leading to the decrease in genetic diversity from the wild progenitor to landraces and from landraces to the modern elite cultivars. we reported that whole genome re-sequencing analysis enhanced our understanding of genetic diversity in wild and cultivated soybeans, and unraveled the processes how this important legume species was domesticated. in present study, the strength of genetic bottlenecks caused by domestication and modern breeding were demonstrated. the continuing reduction of genetic diversity in the cultivated soybean has become a bottleneck for improvement of soybean cultivars. we currently have unprecedented opportunities to exploit genetic diversity in the wild soybean and landraces for sustainable enhancement of soybeans. a set of candidate genes/regions were identified, significantly impacted by selection, for constructing preferred traits underlying soybean domestication and genetic improvement. comparison of candidate domestication and crop improvement-related genes with previous qtl mapping results, as well as their homologs, provides information on potential function of genes under artificial selection. in particular, we found genes related to seed-coat color, growth habit, flowering time and seed size, which had been confirmed as continuously changing from wild soybeans to landraces and then elite cultivars. further analysis is required to identify how variation in these candidate genes affect phenotypes using qtl mapping e.g. in maize, association mapping e.g. in barley, gene expression assays e.g. in sunflower, and gene-knock-out methods. our findings, however, promote development of more efficient approaches to identify the genes underlying domestication-related traits. this study also contribut to construct a large-scale soybean haplotype map and discover important trait related genes using genome-wide association studies. our understanding of the nature of genetic diversity in wild and cultivated soybeans, and the impact of domestication and breeding on genome diversity, will aid future breeding of elite cultivars to improve soybean production and meet the increasing worldwide demands for feed, vegetable oil, soyfood and biofuels. 
artificial selection played an important role in the origin of modern glycine max cultivars from the wild soybean glycine soja. only a small proportion of the whole genomic regions appear to be affected by artificial selection for preferred agricultural traits. given the uniqueness of the soybean germplasm sequenced, this study drew a clear picture of human-mediated evolution of the soybean genomes. we found that artificial selection during domestication led to more pronounced reduction in the genetic diversity of soybean than the switch from landraces to elite cultivars. to elucidate the consequences of artificial selection accompanying the domestication and modern improvement of soybean, new and published whole-genome re-sequencing accessions, which represent wild, domesticated landrace, and chinese elite soybean populations were analyzed. moreover, a set of candidate genes significantly affected by selection underlying soybean domestication and genetic improvement were identified. the selection regions were not distributed randomly or uniformly throughout the genome. instead, clusters of selection hotspots in certain genomic regions were observed.
 artificial selection played an important role in the origin of modern glycine max cultivars from the wild soybean glycine soja. to elucidate the consequences of artificial selection accompanying the domestication and modern improvement of soybean, new and published whole-genome re-sequencing accessions, which represent wild, domesticated landrace, and chinese elite soybean populations were analyzed.  a total of, single nucleotide polymorphisms and insertion/deletions were identified. among the snps detected, % were not described previously. we found that artificial selection during domestication led to more pronounced reduction in the genetic diversity of soybean than the switch from landraces to elite cultivars. only a small proportion of the whole genomic regions appear to be affected by artificial selection for preferred agricultural traits. the selection regions were not distributed randomly or uniformly throughout the genome. instead, clusters of selection hotspots in certain genomic regions were observed. moreover, a set of candidate genes significantly affected by selection underlying soybean domestication and genetic improvement were identified.  given the uniqueness of the soybean germplasm sequenced, this study drew a clear picture of human-mediated evolution of the soybean genomes. the genomic resources and information provided by this study would also facilitate the discovery of genes/loci underlying agronomically important traits. artificial selectionevolutiongenetic diversitypopulation genomicssoybean 