Patent Application: US-72703496-A

Abstract:
the present invention is drawn to a dna encoding a novel ldl receptor - analog . ldl receptors participate in lipoprotein metabolism , which is a critical factor in the onset of arteriosclerosis . the invention provides dna having the nucleotide sequences shown by seq id nos : 1 and 5 , which encode rabbit and human ldl receptor analog proteins having the sequences shown in seq id nos : 3 and 7 respectively .

Description:
the cdna of the present invention may be prepared , for example , by the following process . briefly , the process includes the following steps . ( 1 ) through the use of rabbit ldl receptor cdna as a probe , positive clones are screened out of a rabbit liver cdna library . ( 2 ) recombinant dna is prepared using the separated positive clones , and a cdna fragment is cut out of the resultant recombinant dna through a treatment using a restriction enzyme . the cdna fragment is integrated into a plasmid vector . ( 3 ) host cells are transformed using the obtained cdna recombinant vector to thereby obtain transformant cells of the present invention . the obtained transformant cells are incubated so as to obtain a recombinant vector containing a dna fragment of the present invention . the nucleotide sequence of the dna fragment of the present invention contained in the resultant recombinant vector is determined . ( 4 ) in tissue of a living body , there is detected expression of mrna indicated by the nucleotide sequence of the cdna of the present invention by using rna blot hybridization method . ( 5 ) through use of a rabbit cdna fragment as a probe , positive clones are screened out of a human tissue cdna library , and the nucleotide sequence of the clones is determined . ( 6 ) a recombinant vector for expression is prepared using the cdna of the present invention . through use of the thus - obtained vector , host cells are transformed to thereby obtain the transformants of the present invention . ( 7 ) ligands that are bound to protein expressed by the obtained transformants are detected by ligand blotting . ( 1 ) screening for positive clones from a rabbit liver cdna library : a cdna library may be prepared by the use of mrna obtained from rabbit liver , reverse transcriptase , and a suitable vector , e . g ., commercially available λgt10 vector . a cdna library thus prepared using λgt10 as a vector is subjected to a screening for positive clones by the application of a dna hybridization method employing a cdna probe , to thereby separate positive clones [ sambrook , j ., fritsch , e . f . and maniatis , t . ( 1989 ) in : molecular cloning : a laboratory manual , pp 9 . 47 - 9 . 58 , cold spring harbor laboratory press ]. an exemplary cdna which may be used as a probe is rabbit ldl receptor cdna . positive clones may be detected by autoradiography employing a dna probe labelled with a radioisotope ( 32 p ). recombinant vector λgt10 phage dna is extracted from the isolated positive clones and purified . the resultant purified recombinant vector λgt10 phage dna is digested with a restriction enzyme ecori , to thereby separate a cdna fragment from the vector dna . the obtained cdna fragment is integrated with a plasmid vector for cloning that has been similarly digested with ecori , thereby obtaining a recombinant plasmid vector . an exemplary plasmid vector which may be used is pbluescript ii . ( 3 ) recombinant vector , transformation of host cells using the recombinant vector , and preparation of dna : the obtained cdna recombinant vector is introduced into a variety of host cells that are capable of utilizing the genetic marker possessed by the recombinant vector , to thereby transform the host cells . host cells are not particularly limited , with e . coli being preferred . for example , a variety of variants of the e . coli k12 strain , e . g ., hb - 101 , may be used . in order to introduce the recombinant vector into host cells , a competent cell method may be used in combination with a treatment with calcium . the thus - obtained transformant cells are cultured in a selective medium in accordance with the genetic marker of the vector . the recombinant vector of the present invention is collected from the cultured cells . the dna nucleotide sequence of the cdna contained in the obtained recombinant vector can be determined through use of a dideoxy sequence method [ sanger , f ., nicklen , s . and coulson , a . r . ( 1977 ) proc . natl . acd . sci . u . s . a . 74 , 5463 - 5467 ]. the expression in tissue of mrna , indicated by the nucleotide sequence of the cdna of the present invention , is detected using rna blot hybridization . first , mrna is prepared using rabbit tissue . commercially available oligo ( dt ) cellulose column may be used for the preparation . in order to prepare mrna from human tissue , there may be used a commercially available nylon membrane on which tissue poly ( a ) + rna from a variety of sources is present . an exemplary probe is the rabbit cdna obtained in the above - described step ( 3 ). mrna may be detected by autoradiography employing a dna probe labelled with a radioisotope ( 32 p ). ( 5 ) screening of human tissue cdna library for positive clones , and determination of nucleotide sequence : an exemplary human tissue cdna library which may be used is a commercially available human brain cdna library . screening and nucleotide sequencing of the human brain cdna library may be performed using a fragment of rabbit cdna of the present invention as a probe in a manner similar to that used for the aforementioned rabbit liver cdna library . ( 6 ) preparation of a recombinant vector for expression and transformation of host cells using the recombinant vector for expression : in order to prepare an ldl receptor analog protein through use of cdna of the present invention , the obtained cdna and a vector for expression are first bonded to each other to thereby create a recombinant vector for expression . vectors for expression which may be used for bonding are not particularly limited . for example , pbk - cmv may be used . host cells are transformed using the thus - obtained recombinant vector for expression , to thereby obtain a transformant cell of the present invention . the obtained transformant cell is cultured so as to obtain cells that are capable of expressing the protein of the invention . host cells are not particularly limited . for example , cho cells may be used . in order to introduce the recombinant vector for expression into host cells , a calcium phosphate method may be used . the thus - prepared transformant cells are incubated in a selective medium in accordance with the genetic marker of the vector , so as to express the ldl receptor analog protein of the present invention . after the resultant transformant cells are incubated , the expressed ldl receptor analog protein is solubilized using a solubilizer , e . g ., triton x - 100 , to thereby obtain a membrane protein fraction . the fraction is separated using sds - page , and transferred onto , for example , a nitrocellulose membrane . using a radio - labelled ( 125 i ) lipoprotein as a probe , the analog protein can be detected by autoradiography . exemplary lipoproteins which may be used include β - vldl and ldl . the present invention will next be described in detail by way of example , which should not be construed as limiting the invention . from tissue of the liver of a male japanese white rabbit , intact rna was extracted through a guanidium thiocyanate / cesium chloride method . the obtained intact rna was subjected to an oligo ( dt ) cellulose column method to thereby obtain purified poly ( a ) + rna . cdna was synthesized in accordance with a method of gubler and hoffman [ gubler , u . and hoffman , b . j . ( 1983 ) gene 25 , 263 ]. briefly , cdna was synthesized employing rabbit liver poly ( a ) + rna ( as a template ), a random primer , and moloney murine leukemia virus reverse transcriptase . the synthesized cdna was transformed into double - stranded dna using dna polymerase i , and then subjected to an ecori methylase treatment . by the use of t4 dna polymerase , the dna was blunt - ended . the blunt - ended dna was ligated to phosphorylated ecori linker pd ( ccgaattcgg ) ( seq id no : 8 ) using a t4 dna ligase , and the resultant ligated product was subjected to an additional digestion with ecori . cdna fragments having a size not less than 1 kb were selected by agarose gel electrophoresis , and integrated into the ecori - digested site of λgt10 phage dna using a t4 dna ligase . the phage dna was packaged in vitro , to thereby establish a rabbit liver cdna library . the cdna library ( 1 , 000 , 000 plaques ) prepared in example 1 was subjected to screening using a plaque hybridization method and employing as a probe a segment of the cdna obtained from a ligand binding region , the functional region , of the rabbit ldl receptor . hybridization was performed at 42 ° c . using 5 × ssc , 30 % formamide , 1 % sds , 5 × denhardt &# 39 ; s , and 100 μg / ml salmon sperm dna ( ssdna ), followed by washing with 0 . 3 × ssc / 0 . 1 % sds at 48 ° c . as a result , several positive clones were obtained . these cdna clones were separated by performing this plaque hybridization method in a plurality of times . subsequently , a cdna fragment of each phage was subcloned into a plasmid vector pbluescript ii , and the nucleotide sequence was analyzed using a dideoxy sequence method [ sanger , f ., nicklen , s . and coulson , a . r . ( 1977 ) proc . natl . acd . sci . u . s . a . 74 , 5463 - 5467 ]. based on a putative amino acid sequence , ldl receptors themselves were excluded , and cdna clones having a sequence very similar to that of ldl receptors were identified . using these clones as cdna probes , the cdna library was screened to thereby obtain overlapping two clones . these were employed as new probes and similar procedure was performed , so as to obtain 5 cdna clones . the dna nucleotide sequence determined by these cdna clones are shown as sequence id no . 2 . the total length of the sequence was 6961 bp . in the open reading frame of 6639 bp ( sequence id no . 1 ) which contained a sequence exhibiting high homology with ldl receptors , there existed on the 5 &# 39 ; side an atg codon which was presumably a translation initiating site and a successive highly hydrophobic sequence consisting of about 30 amino acids . accordingly , the obtained cdna was considered to contain the entirety of its length . a putative amino acid sequence is shown as sequence id no . 3 . the protein consisted of 2213 amino acids . comparison of the amino acid sequence of the protein with other amino acid sequence data registered at the genebank , there was a very high similarity to ldl receptors . that is , amino acids 700 - 1 , 100 in the sequence were very similar to the egf precursor homology region of ldl receptors , and amino acids 1 , 100 - 1 , 640 were also very similar to the ligand binding region of ldl receptors . when the amino acid sequence of the subject protein was compared with other lipoprotein receptor lrp , gp330 , and vldl receptors , similarity was not as high as that observed for ldl receptors . on the c - terminal side of the amino acid sequence of the protein , there was found a highly hydrophobic region which was very similar to the transmembrane region of ldl receptors . from liver tissue and brain tissue of a male japanese white rabbit , intact rna was extracted through a guanidium thiocyanate / cesium chloride method . the obtained intact rna was subjected to an oligo ( dt ) cellulose column method to thereby obtain purified poly ( a ) + rna . the poly ( a ) + rna specimens ( 10 μg each ) was modified via a glyoxal method , electrophoresed on 1 % agarose gel , and transferred onto a nylon membrane . for human tissue mrna , commercially available nylon membranes blotted with human tissue poly ( a ) + rna from various sources were used . using as a probe part of a 32 p - labelled rabbit cdna of the present invention , hybridization was performed at 42 ° c . using 50 % ( rabbit ) or 40 % ( human ) formamide , 0 . 1 % sds , 50 mm phosphate buffer , 5 × denhardt &# 39 ; s , 5 × ssc , and 200 μg / ml of ssdna , followed by washing with 0 . 1 × ssc and 0 . 1 % sds at 50 ° c . autoradiography was performed at - 70 ° c . for 2 days in the presence of intensifying screen . as a result , in both rabbit liver tissue and brain tissue , mrna of about 7 kb was detected as well as mrna of about 15 kb which was considered to result from alternative splicing or polyadenylation . the size of the mrna of about 7 kb coincided with that of the rabbit cdna of the present invention . also , in human liver tissue and brain tissue , it was confirmed that mrna having the same size was expressed . screening of human brain cdna library for positive clones and determination of the nucleotide sequence of cdna fragments the human brain cdna library used in this example was a commercially obtained cdna library which was constructed using λgt10 as a vector . using partial cdna of the present invention as a probe , screening of the cdna library ( 300 , 000 plaques ) was performed using a plaque hybridization method . procedures of screening , cloning , and sequencing were as described in example 2 of the present invention . as a result of screening of the human brain cdna library , positive clones containing a dna fragment of about 3 kb were obtained . analysis of the nucleotide sequence of part of the cdna fragment revealed that the fragment was highly homologous to the cdna of the present invention ( sequence id no . 4 ). a human brain cdna library was subjected to screening using fragments of the cdna of the present invention and fragments of the cdna obtained in example 4 as probes . procedures of screening , cloning , and sequencing were as described in example 2 of the present invention . through screening of the human brain cdna library , two positive clones containing cdna fragments of about 6 kb and about 3 kb were obtained . when their nucleotide sequence was analyzed , they were identified to be a cdna clone containing the cdna nucleotide sequence obtained in example 4 and a cdna clone that overlapped therewith . using part of these cdnas as probes , procedures similar to those as described above were performed , to thereby obtain another cdna clone . the dna nucleotide sequence indicated by these cdna clones are shown as sequence id no . 6 . the total length of the sequence was 6 , 843 bp . there was an open reading frame having a size of 6 , 642 bp ( sequence id no . 5 ). a putative amino acid sequence is shown as sequence id no . 7 . the protein consisted of 2 , 214 amino acids . comparison of the amino acid sequence with that of rabbit protein shown by sequence id no . 3 revealed high homology of not less than 94 %. creation of cells that express receptors in the rabbit ldl receptor family : the cdna as shown by sequence id no . 2 was ligated to phosphorylated ecori linker pd ( ccgaattcgg ) ( seq id no : 8 ) by the use of a t4 dna ligase , and the resultant ligated product was digested with ecori . separately , a vector for expression , pbk - cmv was digested with ecori . the aforementioned dna was ligated to the ecori - digested site of the vector using a t4 dna ligase . using the resultant recombinant expression vector in a calcium phosphate method [ chen , c . and h . okayama ( 1987 ) mol . cell . biol . 7 , 2945 - 2752 ], host cells ( cho - 1d1a7 ) were transformed . the resultant transformants were incubated in a ham &# 39 ; s f - 12 selective medium supplemented with 500 μg / ml of g418 , and viable cells were separated as ldl receptor analog protein - expressing cells . the cells were incubated further in the aforementioned medium . ligand analysis of the ldl receptor analog protein by ligand blotting : the obtained ldl receptor analog protein - expressing cells and control cells were suspended in a buffer solution containing 200 mm tris - maleic acid ( ph 6 . 5 ), 2 mm calcium chloride , 0 . 5 mm pmsf , 2 . 5 μm leupeptin , and 1 % triton x - 100 , to thereby solubilize the membrane protein . solubilized membrane protein fractions were obtained through centrifugation , and electrophoresed by a 4 . 5 - 18 % gradient sds - page . thereafter , the protein was transferred onto a nitrocellulose membrane . incubation was performed in a buffer of 50 mm tris - hcl ( ph 8 . 0 ) containing 125 i - labelled β - vldl ( 10 μg / ml ), 2 mm calcium chloride , and 5 % bovine serum albumin . autoradiography was performed at room temperature . a single band of about 250 kda was detected in membrane protein fractions prepared using the present protein - expressing cells . this size coincided well with the molecular weight of 248 kda calculated regarding the amino acid sequence ( sequence id no . 3 ) deduced from the cdna of the present invention . although a similar band was detected for control cells , the expression level was much lower as compared with the case of the present protein - expressing cells . since the protein coded by the cdna of the present invention is considered to be a novel ldl receptor family receptor , it is expected that through analyses of this protein , details of lipoprotein metabolism mediated by the membrane receptor will be elucidated , and pathology of abnormal lipid metabolism which triggers onset and progress of arteriosclerosis will be clarified . __________________________________________________________________________sequence listing ( 1 ) general information :( iii ) number of sequences : 8 ( 2 ) information for seq id no : 1 :( i ) sequence characteristics :( a ) length : 6639 base pairs ( b ) type : nucleic acid ( c ) strandedness : double ( d ) topology : linear ( ii ) molecule type : cdna to mrna ( xi ) sequence description : seq id no : 1 : atggcgacacggagcagcaggagggagtcgcgactccccttcctattcaccctggtcgcg60ctgctgccgcccggggctctctgcgaggtgtggacgcggacactgcacggcggccgcgcg120cccttaccccaggagcggggcttccgcgtggtgcagggcgacccgcgcgagctgcggctg180tgggagcgcggggatgccaggggggcgagccgggcggacgagaagccgctccggaggaga240cggagcgctgccctgcagcccgagcccatcaaggtgtacggacaggtcagcctcaatgat300tcccacaatcagatggtggtgcactgggccggagagaaaagcaacgtgatcgtggccttg360gcccgggacagcctggcgttggccaggcccaggagcagtgatgtgtacgtgtcttatgac420tatggaaaatcattcaataagatttcagagaaattgaacttcggcgcgggaaataacaca480gaggctgtggtggcccagttctaccacagccctgcggacaacaaacggtacatcttcgca540gatgcctacgcccagtatctctggatcacgtttgacttctgcaacaccatccatggcttt600tccatcccgttccgggcagctgatctcctactccacagtaaggcctccaaccttctcctg660ggcttcgacaggtctcaccccaacaagcagctgtggaagtcggatgattttggccagacc720tggatcatgattcaagaacacgtgaagtccttttcttggggaattgatccctatgacaaa780ccaaacaccatctacatcgaacggcacgaaccttctggctactccacggttttccgaagt840acagacttcttccagtcccgggaaaaccaggaagtgatcttggaggaagtgagagacttt900cagcttcgggacaagtacatgtttgctacaaaggtggtgcatctcttgggcagtccactg960cagtcttctgtccagctctgggtctcctttggccggaagcccatgcgggccgcccagttt1020gttacaagacatcctatcaacgaatattacatcgcggatgcctcggaggaccaggtgttt1080gtgtgtgtcagtcacagcaacaaccgcaccaacctctacatctcggaggcagagggcttg1140aagttctctctgtccctggagaacgtgctctactacaccccgggaggggccggcagtgac1200accttggtgaggtactttgcaaatgaaccgtttgctgacttccatcgtgtggaagggttg1260cagggagtctacattgctactctgattaatggttctatgaatgaggagaacatgagatct1320gtcatcacctttgacaaagggggcacctgggaatttctgcaggctccagccttcacgggg1380tatggagagaaaatcaactgtgagctgtccgagggctgttccctccacctggcccagcgc1440ctcagccagctgctcaacctccagctccggaggatgcccatcctgtccaaggagtcggcg1500cctggcctcatcattgccacgggctcagtgggaaagaacttggctagcaagacaaacgtg1560tacatctctagcagtgctggagccaggtggcgagaggcacttcctggacctcactactat1620acatggggagaccatggcggcatcatcatggccattgcccaaggcatggaaaccaacgaa1680ctgaagtacagtaccaacgaaggggagacctggaaagccttcaccttctctgagaagccc1740gtgtttgtgtatgggctcctcacggaacccggcgagaagagcacggtcttcaccatcttt1800ggctccaacaaggagaacgtgcacagctggctcatcctccaggtcaatgccacagacgcc1860ctgggggttccttgcacagagaacgactacaagctctggtcaccatctgatgagcggggg1920aatgagtgtttgcttggacacaagactgttttcaaacggaggaccccgcacgccacatgc1980tttaacggagaagactttgacaggccggtggttgtgtccaactgctcctgcacccgggag2040gactatgagtgtgactttggcttccggatgagtgaagacttggcattagaggtgtgtgtt2100ccagatccaggattttctggaaagtcctcccctccagtgccttgtcccgtgggctctacg2160tacaggcgatcaagaggctaccggaagatttctggggacacctgtagtggaggagatgtt2220gaggcacggctagaaggagagctggtcccctgtcccctggcagaagagaacgagttcatc2280ctgtacgccacgcgcaagtccatccaccgctatgacctggcttccggaaccacggagcag2340ttgcccctcactgggttgcgggcagcagtggccctggactttgactatgagcacaactgc2400ctgtattggtctgacctggccttggacgtcatccagcgcctctgtttgaacgggagtaca2460ggacaagaggtgatcatcaactctgacctggagacggtagaagctttggcttttgaaccc2520ctcagccaattactttactgggtggacgcaggctttaaaaagatcgaggtagccaatcca2580gatggtgacttccgactcaccgtcgtcaattcctcggtgctggatcggccccgggccctg2640gtccttgtgccccaagaagggatcatgttctggaccgactggggagacctgaagcctggg2700atttatcggagcaacatggacggatctgccgcctatcgcctcgtgtcggaggatgtgaag2760tggcccaatggcatttccgtggacgatcagtggatctactggacggatgcctacctggac2820tgcattgagcgcatcacgttcagcggccagcagcgctccgtcatcctggacagactcccg2880cacccctatgccattgctgtctttaagaatgagatttactgggatgactggtcacagctc2940agcatattccgagcttctaagtacagcgggtcccagatggagattctggccagccagctc3000acggggctgatggacatgaagatcttctacaaggggaagaacacaggaagcaatgcgtgt3060gtacccaggccgtgcagcctgctgtgcctgcccagagccaacaacagcaaaagctgcagg3120tgtccagatggcgtggccagcagtgtcctcccttccggggacctgatgtgtgactgccct3180aagggctacgagctgaagaacaacacgtgtgtcaaagaagaagacacctgtctgcgcaac3240cagtaccgctgcagcaacgggaactgcatcaacagcatctggtggtgcgatttcgacaac3300gactgcggagacatgagcgacgagaagaactgccctaccaccatctgcgacctggacacc3360cagttccgttgccaggagtctgggacgtgcatcccgctctcctacaaatgtgacctcgag3420gatgactgtggggacaacagtgacgaaaggcactgtgaaatgcaccagtgccggagcgac3480gaatacaactgcagctcgggcatgtgcatccgctcctcctgggtgtgcgacggggacaac3540gactgcagggactggtccgacgaggccaactgcacagccatctatcacacctgtgaggcc3600tccaacttccagtgccgcaacgggcactgcatcccccagcggtgggcgtgtgacggcgac3660gccgactgccaggatggctctgatgaggatccagccaactgtgagaagaagtgcaacggc3720ttccgctgcccgaacggcacctgcattccctccaccaagcactgtgacggcctgcacgat3780tgctcggacggctccgacgagcagcactgcgagcccctgtgtacacggttcatggacttc3840gtgtgtaagaaccgccagcagtgcctcttccactccatggtgtgcgatgggatcatccag3900tgccgtgacggctccgacgaggacccagcctttgcaggatgctcccgagaccccgagttc3960cacaaggtgtgcgatgagttcggcttccagtgtcagaacggcgtgtgcatcagcttgatc4020tggaagtgcgacgggatggatgactgcggggactactccgacgaggccaactgtgaaaac4080cccacagaagcccccaactgctcccgctacttccagttccggtgtgacaatggccactgc4140atccccaacaggtggaagtgtgacagggagaatgactgtggggactggtccgacgagaag4200gactgtggagattcacatgtacttccgtctacgactcctgcaccctccacgtgtctgccc4260aattactaccgctgcggcgggggggcctgcgtgatagacacgtgggtttgtgacgggtac4320cgagattgcgcagatggatccgacgaggaagcctgcccctcgctccccaatgtcactgcc4380acctcctccccctcccagcctggacgatgcgaccgatttgagtttgagtgccaccagcca4440aagaagtgcatccctaactggagacgctgtgacggccatcaggattgccaggatggccag4500gacgaggccaactgccccactcacagcaccttgacctgcatgagctgggagttcaagtgt4560gaggatggcgaggcctgcatcgtgctgtcagaacgctgcgacggcttcctggactgctca4620gatgagagcgacgagaaggcctgcagtgatgagttaactgtatacaaagtacagaatctt4680cagtggacagctgacttctctgggaatgtcactttgacctggatgcggcccaaaaaaatg4740ccctctgctgcttgtgtatacaacgtgtactatagagttgttggagagagcatatggaag4800actctggagactcacagcaataagacaaacactgtattaaaagtgttgaaaccagatacc4860acctaccaggttaaagtgcaggttcagtgcctgagcaaggtgcacaacaccaatgacttt4920gtgaccttgagaactccagagggattgccagacgcccctcagaacctccagctgtcgctc4980cacggggaagaggaaggtgtgattgtgggccactggagccctcccacccacacccacggc5040ctcattcgcgaatacattgtagagtatagcaggagtggttccaaggtgtggacttcagaa5100agggctgctagtaactttacagaaataaagaacttgttggtcaacaccctgtacaccgtc5160agagtggctgcggtgacgagtcgtgggataggaaactggagcgattccaaatccattacc5220accgtgaaaggaaaagcgatcccgccaccaaatatccacattgacaactacgatgaaaat5280tccctgagttttaccctgaccgtggatgggaacatcaaggtgaatggctatgtggtgaac5340cttttctgggcatttgacacccacaaacaagagaagaaaaccatgaacttccaagggagc5400tcagtgtcccacaaagttggcaatctgacagcacagacggcctatgagatttccgcctgg5460gccaagactgacttgggcgatagtcctctgtcatttgagcatgtcacgaccagaggggtt5520cgcccacctgctcctagcctcaaggccagggctatcaatcagactgcagtggaatgcacc5580tggacaggccccaggaatgtggtgtatggcattttctatgccacatccttcctggacctc5640taccgcaacccaagcagcctgaccacgccgctgcacaacgcaaccgtgctcgtcggtaag5700gatgagcagtatctgtttctggtccgggtggtgatgccctaccaagggccgtcctcggac5760tacgtggtcgtgaagatgatcccggacagcaggcttcctccccggcacctgcatgccgtt5820cacaccggcaagacctcggccgtcatcaagtgggagtcgccctacgactctcctgaccag5880gacctgttctatgcgatcgcagttaaagatctgatacgaaagacggaccggagctacaaa5940gtcaagtcccgcaacagcaccgtggagtacaccctgagcaagctggagcccggagggaaa6000taccacgtcattgtgcagctggggaacatgagcaaagatgccagtgtgaagatcaccacc6060gtttcgttatcggcacccgatgccttaaaaatcataacagaaaatgaccacgtccttctc6120ttctggaaaagtctagctctaaaggaaaagtattttaacgaaagcaggggctacgagata6180cacatgtttgatagcgccatgaatatcaccgcataccttgggaatactactgacaatttc6240tttaaaatttccaacctgaagatgggtcacaattacacattcacggtccaggcacgatgc6300cttttgggcagccagatctgcggggagcctgccgtgctactgtatgatgagctggggtct6360ggtggcgatgcgtcggcgatgcaggctgccaggtctactgatgtcgccgccgtggtggtg6420cccatcctgtttctgatactgctgagcctgggggtcgggtttgccatcctgtacacgaag6480catcggaggctgcagagcagcttcaccgccttcgccaacagccactacagctccagactc6540ggctccgccatcttctcctctggggatgacttgggggaggatgatgaagatgctcctatg6600atcactggattttcggacgacgtccccatggtgatagcc6639 ( 2 ) information for seq id no : 2 :( i ) sequence characteristics :( a ) length : 6961 base pairs ( b ) type : nucleic acid ( c ) strandedness : double ( d ) topology : linear ( ii ) molecule type : cdna to mrna ( ix ) feature :( a ) name / key : cds ( b ) location : 178 .. 6819 ( d ) other information : / note =&# 34 ; identification method : s &# 34 ;( ix ) feature :( a ) name / key : sig_peptide ( b ) location : 178 .. 261 ( d ) other information : / note =&# 34 ; identification method : s &# 34 ;( ix ) feature :( a ) name / key : misc_feature ( b ) location : 262 .. 6816 ( d ) other information : / function =&# 34 ; nucleotides 262 - 6816encode the mature peptide &# 34 ;/ note =&# 34 ; identification method : s &# 34 ;( xi ) sequence description : seq id no : 2 : 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* 2210gtaaatattttatttgataaagatagttgatggtttattttaaaagatgcactttgagtt6909gcaatatgttatttttatatgggccaaaaacaaaagcaaaaaaaaaaaaaaa6961 ( 2 ) information for seq id no : 3 :( i ) sequence characteristics :( a ) length : 2213 amino acids ( b ) type : amino acid ( d ) topology : linear ( ii ) molecule type : protein ( xi ) sequence description : seq id no : 3 : 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6513701375asnglyhiscysileproasnargtrplyscysasparggluasnasp138013851390cysglyasptrpseraspglulysaspcysglyaspserhisvalleu139514001405proserthrthrproalaproserthrcysleuproasntyrtyrarg141014151420cysglyglyglyalacysvalileaspthrtrpvalcysaspglytyr1425143014351440argaspcysalaaspglyseraspgluglualacysproserleupro144514501455asnvalthralathrserserproserglnproglyargcysasparg146014651470pheglupheglucyshisglnprolyslyscysileproasntrparg147514801485argcysaspglyhisglnaspcysglnaspglyglnaspglualaasn149014951500cysprothrhisserthrleuthrcysmetsertrpgluphelyscys1505151015151520gluaspglyglualacysilevalleusergluargcysaspglyphe152515301535leuaspcysseraspgluseraspglulysalacysseraspgluleu154015451550thrvaltyrlysvalglnasnleuglntrpthralaaspphesergly155515601565asnvalthrleuthrtrpmetargprolyslysmetproseralaala157015751580cysvaltyrasnvaltyrtyrargvalvalglygluseriletrplys1585159015951600thrleugluthrhisserasnlysthrasnthrvalleulysvalleu160516101615lysproaspthrthrtyrglnvallysvalglnvalglncysleuser162016251630lysvalhisasnthrasnaspphevalthrleuargthrproglugly163516401645leuproaspalaproglnasnleuglnleuserleuhisglygluglu165016551660gluglyvalilevalglyhistrpserproprothrhisthrhisgly1665167016751680leuileargglutyrilevalglutyrserargserglyserlysval168516901695trpthrsergluargalaalaserasnphethrgluilelysasnleu170017051710leuvalasnthrleutyrthrvalargvalalaalavalthrserarg171517201725glyileglyasntrpseraspserlysserilethrthrvallysgly173017351740lysalaileproproproasnilehisileaspasntyraspgluasn1745175017551760serleuserphethrleuthrvalaspglyasnilelysvalasngly176517701775tyrvalvalasnleuphetrpalapheaspthrhislysglnglulys178017851790lysthrmetasnpheglnglyserservalserhislysvalglyasn179518001805leuthralaglnthralatyrgluileseralatrpalalysthrasp181018151820leuglyaspserproleuserphegluhisvalthrthrargglyval1825183018351840argproproalaproserleulysalaargalaileasnglnthrala184518501855valglucysthrtrpthrglyproargasnvalvaltyrglyilephe186018651870tyralathrserpheleuaspleutyrargasnproserserleuthr187518801885thrproleuhisasnalathrvalleuvalglylysaspgluglntyr189018951900leupheleuvalargvalvalmetprotyrglnglyproserserasp1905191019151920tyrvalvalvallysmetileproaspserargleuproproarghis192519301935leuhisalavalhisthrglylysthrseralavalilelystrpglu194019451950serprotyraspserproaspglnaspleuphetyralailealaval195519601965lysaspleuilearglysthraspargsertyrlysvallysserarg197019751980asnserthrvalglutyrthrleuserlysleugluproglyglylys1985199019952000tyrhisvalilevalglnleuglyasnmetserlysaspalaserval200520102015lysilethrthrvalserleuseralaproaspalaleulysileile202020252030thrgluasnasphisvalleuleuphetrplysserleualaleulys203520402045glulystyrpheasngluserargglytyrgluilehismetpheasp205020552060seralametasnilethralatyrleuglyasnthrthraspasnphe2065207020752080phelysileserasnleulysmetglyhisasntyrthrphethrval208520902095glnalaargcysleuleuglyserglnilecysglygluproalaval210021052110leuleutyraspgluleuglyserglyglyaspalaseralametgln211521202125alaalaargserthraspvalalaalavalvalvalproileleuphe213021352140leuileleuleuserleuglyvalglyphealaileleutyrthrlys2145215021552160hisargargleuglnserserphethralaphealaasnserhistyr216521702175serserargleuglyseralailepheserserglyaspaspleugly218021852190gluaspaspgluaspalaprometilethrglypheseraspaspval219522002205prometvalileala2210 ( 2 ) information for seq id no : 4 :( i ) sequence characteristics :( a ) length : 300 base pairs ( b ) type : nucleic acid ( c ) strandedness : double ( d ) topology : linear ( ii ) molecule type : cdna to mrna ( xi ) sequence description : seq id no : 4 : atatccacattgacagctatggtgaaaattatctaagcttcaccctgaccatggagagtg60atatcaaggtgaatggctatgtggtgaaccttttctgggcatttgacacccacaagcaag120agaggagaactttgaacttccgaggaagcatattgtcacacaaagttggcaatctgacag180ctcatacatcctatgagatttctgcctgggccaagactgacttgggggatagccctctgg240catttgagcatgttatgaccagaggggttcgcccacctgcacctagcctcaaggccaaag300 ( 2 ) information for seq id no : 5 :( i ) sequence characteristics :( a ) length : 6642 base pairs ( b ) type : nucleic acid ( c ) strandedness : double ( d ) topology : linear ( ii ) molecule type : cdna to mrna ( xi ) sequence description : seq id no : 5 : atggcgacacggagcagcaggagggagtcgcgactcccgttcctattcaccctggtcgca60ctgctgccgcccggagctctctgcgaagtctggacgcagaggctgcacggcggcagcgcg120cccttgccccaggaccggggcttcctcgtggtgcagggcgacccgcgcgagctgcggctg180tgggcgcgcggggatgccaggggggcgagccgcgcggacgagaagccgctccggaggaaa240cggagcgctgccctgcagcccgagcccatcaaggtgtacggacaggttagtctgaatgat300tcccacaatcagatggtggtgcactgggctggagagaaaagcaacgtgatcgtggccttg360gcccgagatagcctggcattggcgaggcccaagagcagtgatgtgtacgtgtcttacgac420tatggaaaatcattcaagaaaatttcagacaagttaaactttggcttgggaaataggagt480gaagctgttatcgcccagttctaccacagccctgcggacaacaagcggtacatctttgca540gacgcttatgcccagtacctctggatcacgtttgacttctgcaacactcttcaaggcttt600tccatcccatttcgggcagctgatctcctcctacacagtaaggcctccaaccttctcttg660ggctttgacaggtcccaccccaacaagcagctgtggaagtcagatgactttggccagacc720tggatcatgattcaggaacatgtcaagtccttttcttggggaattgatccctatgacaaa780ccaaataccatctacattgaacgacacgaaccctctggctactccactgtcttccgaagt840acagatttcttccagtcccgggaaaaccaggaagtgatccttgaggaagtgagagatttt900cagcttcgggacaagtacatgtttgctacaaaggtggtgcatctcttgggcagtgaacag960cagtcttctgtccagctctgggtctcctttggccggaagcccatgagagcagcccagttt1020gtcacaagacatcctattaatgaatattacatcgcagatgcctccgaggaccaggtgttt1080gtgtgtgtcagccacagtaacaaccgcaccaatttatacatctcagaggcagaggggctg1140aagttctccctgtccttggagaacgtgctctattacagcccaggaggggccggcagtgac1200accttggtgaggtattttgcaaatgaaccatttgctgacttccaccgagtggaaggattg1260caaggagtctacattgctactctgattaatggttctatgaatgaggagaacatgagatcg1320gtcatcacctttgacaaagggggaacctgggagtttcttcaggctccagccttcacggga1380tatggagagaaaatcaattgtgagctttcccagggctgttcccttcatctggctcagcgc1440ctcagtcagctcctcaacctccagctccggagaatgcccatcctgtccaaggagtcggct1500ccaggcctcatcatcgccactggctcagtgggaaagaacttggctagcaagacaaacgtg1560tacatctctagcagtgctggagccaggtggcgagaggcacttcctggacctcactactac1620acatggggagaccacggcggaatcatcacggccattgcccagggcatggaaaccaacgag1680ctaaaatacagtaccaatgaaggggagacctggaaaacattcatcttctctgagaagcca1740gtgtttgtgtatggcctcctcacagaacctggggagaagagcactgtcttcaccatcttt1800ggctcgaacaaagagaatgtccacagctggctgatcctccaggtcaatgccacggatgcc1860ttgggagttccctgcacagagaatgactacaagctgtggtcaccatctgatgagcggggg1920aatgagtgtttgctgggacacaagactgttttcaaacggcggaccccccatgccacatgc1980ttcaatggagaggactttgacaggccggtggtcgtgtccaactgctcctgcacccgggag2040gactatgagtgtgacttcggtttcaagatgagtgaagatttgtcattagaggtttgtgtt2100ccagatccggaattttctggaaagtcatactcccctcctgtgccttgccctgtgggttct2160acttacaggagaacgagaggctaccggaagatttctggggacacttgtagcggaggagat2220gttgaagcgcgactggaaggagagctggtcccctgtcccctggcagaagagaacgagttc2280attctgtatgctgtgaggaaatccatctaccgctatgacctggcctcgggagccaccgag2340cagttgcctctcaccgggctacgggcagcagtggccctggactttgactatgagcacaac2400tgtttgtattggtccgacctggccttggacgtcatccagcgcctctgtttgaatggaagc2460acagggcaagaggtgatcatcaattctggcctggagacagtagaagctttggcttttgaa2520cccctcagccagctgctttactgggtagatgcaggcttcaaaaagattgaggtagctaat2580ccagatggcgacttccgactcacaatcgtcaattcctctgtgcttgatcgtcccagggct2640ctggtcctcgtgccccaagagggggtgatgttctggacagactggggagacctgaagcct2700gggatttatcggagcaatatggatggttctgctgcctatcacctggtgtctgaggatgtg2760aagtggcccaatggcatctctgtggacgaccagtggatttactggacggatgcctacctg2820gagtgcatagagcggatcacgttcagtggccagcagcgctctgtcattctggacaacctc2880ccgcacccctatgccattgctgtctttaagaatgaaatctactgggatgactggtcacag2940ctcagcatattccgagcttccaaatacagtgggtcccagatggagattctggcaaaccag3000ctcacggggctcatggacatgaagattttctacaaggggaagaacactggaagcaatgcc3060tgtgtgcccaggccatgcagcctgctgtgcctgcccaaggccaacaacagtagaagctgc3120aggtgtccagaggatgtgtccagcagtgtgcttccatcaggggacctgatgtgtgactgc3180cctcagggctatcagctcaagaacaatacctgtgtcaaagaagagaacacctgtcttcgc3240aaccagtatcgctgcagcaacgggaactgtatcaacagcatttggtggtgtgactttgac3300aacgactgtggagacatgagcgatgagagaaactgccctaccaccatctgtgacctggac3360acccagtttcgttgccaggagtctgggacttgtatcccactgtcctataaatgtgacctt3420gaggatgactgtggagacaacagtgatgaaagtcattgtgaaatgcaccagtgccggagt3480gacgagtacaactgcagttccggcatgtgcatccgctcctcctgggtatgtgacggggac3540aacgactgcagggactggtctgatgaagccaactgtaccgccatctatcacacctgtgag3600gcctccaacttccagtgccgaaacgggcactgcatcccccagcggtgggcgtgtgacggg3660gatacggactgccaggatggttccgatgaggatccagtcaactgtgagaagaagtgcaat3720ggattccgctgcccaaacggcacttgcatcccatccagcaaacattgtgatggtctgcgt3780gattgctctgatggctccgatgaacagcactgcgagcccctctgtacgcacttcatggac3840tttgtgtgtaagaaccgccagcagtgcctgttccactccatggtctgtgacggaatcatc3900cagtgccgcgacgggtccgatgaggatgcggcgtttgcaggatgctcccaagatcctgag3960ttccacaaggtatgtgatgagttcggtttccagtgtcagaatggagtgtgcatcagtttg4020atttggaagtgcgacgggatggatgattgcggcgattattctgatgaagccaactgcgaa4080aaccccacagaagccccaaactgctcccgctacttccagtttcggtgtgagaatggccac4140tgcatccccaacagatggaaatgtgacagggagaacgactgtggggactggtctgatgag4200aaggattgtggagattcacatattcttcccttctcgactcctgggccctccacgtgtctg4260cccaattactaccgctgcagcagtgggacctgcgtgatggacacctgggtgtgcgacggg4320taccgagattgtgcagatggctctgacgaggaagcctgccccttgcttgcaaacgtcact4380gctgcctccactcccacccaacttgggcgatgtgaccgatttgagttcgaatgccaccaa4440ccgaagacgtgtattcccaactggaagcgctgtgacggccaccaagattgccaggatggc4500cgggacgaggccaattgccccacacacagcaccttgacttgcatgagcagggagttccag4560tgcgaggacggggaggcctgcattgtgctctcggagcgctgcgacggcttcctggactgc4620tcggacgagagcgatgaaaaggcctgcagtgatgagttgactgtgtacaaagtacagaat4680cttcagtggacagctgacttctctggggatgtgactttgacctggatgaggcccaaaaaa4740atgccctctgcatcttgtgtatataatgtctactacagggtggttggagagagcatatgg4800aagactctggagacccacagcaataagacaaacactgtattaaaagtcttgaaaccagat4860accacgtatcaggttaaagtacaggttcagtgtctcagcaaggcacacaacaccaatgac4920tttgtgaccctgaggaccccagagggattgccagatgcccctcgaaatctccagctgtca4980ctccccagggaagcagaaggtgtgattgtaggccactgggctcctcccatccacacccat5040ggcctcatccgtgagtacattgtagaatacagcaggagtggttccaagatgtgggcctcc5100cagagggctgctagtaactttacagaaatcaagaacttattggtcaacactctatacacc5160gtcagagtggctgcggtgactagtcgtggaataggaaactggagcgattctaaatccatt5220accaccataaaaggaaaagtgatcccaccaccagatatccacattgacagctatggtgaa5280aattatctaagcttcaccctgaccatggagagtgatatcaaggtgaatggctatgtggtg5340aaccttttctgggcatttgacacccacaagcaagagaggagaactttgaacttccgagga5400agcatattgtcacacaaagttggcaatctgacagctcatacatcctatgagatttctgcc5460tgggccaagactgacttgggggatagccctctggcatttgagcatgttatgaccagaggg5520gttcgcccacctgcacctagcctcaaggccaaagccatcaaccagactgcagtggaatgt5580acctggaccggcccccggaatgtggtttatggtattttctatgccacgtcctttcttgac5640ctctatcgcaacccgaagagcttgactacttcactccacaacaagacggtcattgtcagt5700aaggatgagcagtatttgtttctggtccgtgtagtggtaccctaccaggggccatcctct5760gactacgttgtagtgaagatgatcccggacagcaggcttccaccccgtcacctgcatgtg5820gttcatacgggcaaaacctccgtggtcatcaagtgggaatcaccgtatgactctcctgac5880caggacttgttgtatgcaattgcagtcaaagatctcataagaaagactgacaggagctac5940aaagtaaaatcccgtaacagcactgtggaatacacccttaacaagttggagcctggcggg6000aaataccacatcattgtccaactggggaacatgagcaaagattccagcataaaaattacc6060acagtttcattatcagcacctgatgccttaaaaatcataacagaaaatgatcatgttctt6120ctgttttggaaaagcctggctttaaaggaaaagcattttaatgaaagcaggggctatgag6180atacacatgtttgatagtgccatgaatatcacagcttaccttgggaatactactgacaat6240ttctttaaaatttccaacctgaagatgggtcataattacacgttcaccgtccaagcaaga6300tgcctttttggcaaccagatctgtggggagcctgccatcctgctgtacgatgagctgggg6360tctggtgcagatgcatctgcaacgcaggctgccagatctacggatgttgctgctgtggtg6420gtgcccatcttattcctgatactgctgagcctgggggtggggtttgccatcctgtacacg6480aagcaccggaggctgcagagcagcttcaccgccttcgccaacagccactacagctccagg6540ctggggtccgcaatcttctcctctggggatgacctgggggaagatgatgaagatgcccct6600atgataactggattttcagatgacgtccccatggtgatagcc6642 ( 2 ) information for seq id no : 6 :( i ) sequence characteristics :( a ) length : 6843 base pairs ( b ) type : nucleic acid ( c ) strandedness : double ( d ) topology : linear ( ii ) molecule type : cdna to mrna ( ix ) feature :( a ) name / key : cds ( b ) location : 81 .. 6725 ( d ) other information : / note =&# 34 ; identification method : s &# 34 ;( ix ) feature :( a ) name / key : sig_peptide ( b ) location : 81 .. 164 ( d ) other information : / note =&# 34 ; identification method : s &# 34 ;( ix ) feature :( a ) name / key : misc_feature ( b ) location : 165 .. 6722 ( d ) other information : / function =&# 34 ; nucleotides 165 - 6722encode the mature peptide &# 34 ;/ note =&# 34 ; identification method : s &# 34 ;( xi ) sequence description : seq id no : 6 : 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] 16651670cctcccatccacacccatggcctcatccgtgagtacattgtagaatac5150proproilehisthrhisglyleuileargglutyrilevalglutyr1675168016851690agcaggagtggttccaagatgtgggcctcccagagggctgctagtaac5198serargserglyserlysmettrpalaserglnargalaalaserasn169517001705tttacagaaatcaagaacttattggtcaacactctatacaccgtcaga5246phethrgluilelysasnleuleuvalasnthrleutyrthrvalarg171017151720gtggctgcggtgactagtcgtggaataggaaactggagcgattctaaa5294valalaalavalthrserargglyileglyasntrpseraspserlys172517301735tccattaccaccataaaaggaaaagtgatcccaccaccagatatccac5342serilethrthrilelysglylysvalileproproproaspilehis174017451750attgacagctatggtgaaaattatctaagcttcaccctgaccatggag5390ileaspsertyrglygluasntyrleuserphethrleuthrmetglu1755176017651770agtgatatcaaggtgaatggctatgtggtgaaccttttctgggcattt5438seraspilelysvalasnglytyrvalvalasnleuphetrpalaphe177517801785gacacccacaagcaagagaggagaactttgaacttccgaggaagcata5486aspthrhislysglngluargargthrleuasnpheargglyserile179017951800ttgtcacacaaagttggcaatctgacagctcatacatcctatgagatt5534leuserhislysvalglyasnleuthralahisthrsertyrgluile180518101815tctgcctgggccaagactgacttgggggatagccctctggcatttgag5582seralatrpalalysthraspleuglyaspserproleualapheglu182018251830catgttatgaccagaggggttcgcccacctgcacctagcctcaaggcc5630hisvalmetthrargglyvalargproproalaproserleulysala1835184018451850aaagccatcaaccagactgcagtggaatgtacctggaccggcccccgg5678lysalaileasnglnthralavalglucysthrtrpthrglyproarg185518601865aatgtggtttatggtattttctatgccacgtcctttcttgacctctat5726asnvalvaltyrglyilephetyralathrserpheleuaspleutyr187018751880cgcaacccgaagagcttgactacttcactccacaacaagacggtcatt5774argasnprolysserleuthrthrserleuhisasnlysthrvalile188518901895gtcagtaaggatgagcagtatttgtttctggtccgtgtagtggtaccc5822valserlysaspgluglntyrleupheleuvalargvalvalvalpro190019051910taccaggggccatcctctgactacgttgtagtgaagatgatcccggac5870tyrglnglyproserserasptyrvalvalvallysmetileproasp1915192019251930agcaggcttccaccccgtcacctgcatgtggttcatacgggcaaaacc5918serargleuproproarghisleuhisvalvalhisthrglylysthr193519401945tccgtggtcatcaagtgggaatcaccgtatgactctcctgaccaggac5966servalvalilelystrpgluserprotyraspserproaspglnasp195019551960ttgttgtatgcaattgcagtcaaagatctcataagaaagactgacagg6014leuleutyralailealavallysaspleuilearglysthrasparg196519701975agctacaaagtaaaatcccgtaacagcactgtggaatacacccttaac6062sertyrlysvallysserargasnserthrvalglutyrthrleuasn198019851990aagttggagcctggcgggaaataccacatcattgtccaactggggaac6110lysleugluproglyglylystyrhisileilevalglnleuglyasn1995200020052010atgagcaaagattccagcataaaaattaccacagtttcattatcagca6158metserlysaspserserilelysilethrthrvalserleuserala201520202025cctgatgccttaaaaatcataacagaaaatgatcatgttcttctgttt6206proaspalaleulysileilethrgluasnasphisvalleuleuphe203020352040tggaaaagcctggctttaaaggaaaagcattttaatgaaagcaggggc6254trplysserleualaleulysglulyshispheasngluserarggly204520502055tatgagatacacatgtttgatagtgccatgaatatcacagcttacctt6302tyrgluilehismetpheaspseralametasnilethralatyrleu206020652070gggaatactactgacaatttctttaaaatttccaacctgaagatgggt6350glyasnthrthraspasnphephelysileserasnleulysmetgly2075208020852090cataattacacgttcaccgtccaagcaagatgcctttttggcaaccag6398hisasntyrthrphethrvalglnalaargcysleupheglyasngln209521002105atctgtggggagcctgccatcctgctgtacgatgagctggggtctggt6446ilecysglygluproalaileleuleutyraspgluleuglysergly211021152120gcagatgcatctgcaacgcaggctgccagatctacggatgttgctgct6494alaaspalaseralathrglnalaalaargserthraspvalalaala212521302135gtggtggtgcccatcttattcctgatactgctgagcctgggggtgggg6542valvalvalproileleupheleuileleuleuserleuglyvalgly214021452150tttgccatcctgtacacgaagcaccggaggctgcagagcagcttcacc6590phealaileleutyrthrlyshisargargleuglnserserphethr2155216021652170gccttcgccaacagccactacagctccaggctggggtccgcaatcttc6638alaphealaasnserhistyrserserargleuglyseralailephe217521802185tcctctggggatgacctgggggaagatgatgaagatgcccctatgata6686serserglyaspaspleuglygluaspaspgluaspalaprometile219021952200actggattttcagatgacgtccccatggtgatagcctgaaagagctttc6735thrglypheseraspaspvalprometvalileala * 22052210ctcactagaaaccaaatggtgtaaatattttatttgataaagatagttgatggtttattt6795taaaagatgcactttgagttgcaatatgttatttttatatgggccaaa6843 ( 2 ) information for seq id no : 7 :( i ) sequence characteristics :( a ) length : 2214 amino acids ( b ) type : amino acid ( d ) topology : linear ( ii ) molecule type : protein ( xi ) sequence description : seq id no : 7 : 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( 2 ) information for seq id no : 8 :( i ) sequence characteristics :( a ) length : 10 base pairs ( b ) type : nucleic acid ( c ) strandedness : single ( d ) topology : linear ( ii ) molecule type : other nucleic acid ( a ) description : / desc =&# 34 ; synthetic dna linker &# 34 ;( xi ) sequence description : seq id no : 8 : ccgaattcgg10__________________________________________________________________________