| 质粒名称: | G10 Cas9 MDC123 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 59187 |
| 存储实验室: | Robert Stupar |
|
载体骨架基本信息 |
|
| 载体名称: | pMDC123 |
| 载体出品公司: | Mark D Curtis |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | 10213 bp |
| 完整质粒大小: | 13191 bp |
| 载体修饰: | 添加了G10启动子而不是2X35S启动子，并添加了具有N和C末端核定位信号的甘氨酸最大密码子优化的Cas9。 |
| 载体类型: | 植物表达，CRISPR |
| 筛选标记: | Basta |
| 高拷贝/低拷贝: | -- |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | Glycine Max codon optimized Cas9，Cas9 |
| 物种来源: | Glycine Max |
| 插入大小: | 4140 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | G10 |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | SalI (未破坏) |
| 3' 克隆位点: | PacI (未破坏) |
| 5' 测序引物 | M13_forward20_primer |
| 3' 测序引物 | M13_reverse_primer |
|
文献引用方法 |
|
| 材料和方法部分: | G10 Cas9 MDC123 was a gift from Robert Stupar (Addgene plasmid # 59187 ; http://n2t.net/addgene:59187 ; RRID:Addgene_59187) |
| 参考文献部分: | CRISPR/Cas mutagenesis of soybean and Medicago truncatula using a new web-tool and a modified Cas9 enzyme. Michno JM, Wang X, Liu J, Curtin SJ, Kono TJ, Stupar RM. GM Crops Food. 2015;6(4):243-52. doi: 10.1080/21645698.2015.1106063. 10.1080/21645698.2015.1106063 PubMed 26479970 |
买家导航
载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
载体序列
LOCUS G10 Cas9 MDC123 13204 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION G10 promoting Glycing Max codon optimized Cas9 (N and C terminus
NLS) with bar plant selection.
ACCESSION .
VERSION .
KEYWORDS G10 Cas9 MDC123
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 13204)
AUTHORS Michno JM, Wang X, Liu J, Curtin SJ, Kono TJ, Stupar RM
TITLE CRISPR/Cas mutagenesis of soybean and Medicago truncatula using a
new web-tool and a modified Cas9 enzyme.
JOURNAL GM Crops Food. 2015;6(4):243-52. doi: 10.1080/21645698.2015.1106063.
PUBMED 26479970
REFERENCE 2 (bases 1 to 13204)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..13204
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_feature 5..145
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(71..90)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 231..253
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
rep_origin 489..683
/label=pVS1 oriV
/note="origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
CDS complement(749..1822)
/codon_start=1
/product="replication protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
/label=pVS1 RepA
/translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
LVMRYRNLIEGEASAGS"
CDS complement(2251..2880)
/codon_start=1
/product="stability protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
/label=pVS1 StaA
/translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
misc_feature 4181..4205
/label=RB T-DNA repeat
/note="right border repeat from nopaline C58 T-DNA"
primer_bind 4393..4415
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 4407..4424
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 4408..4424
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
promoter 4689..4735
/label=minimal CaMV 35S promoter
/note="minimal 35S promoter from cauliflower mosaic virus"
CDS 4761..4781
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 4788..8891
/codon_start=1
/product="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
/label=Cas9
/note="generates RNA-guided double strand breaks in DNA"
/translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
RBS 8804..8812
/label=Shine-Dalgarno sequence
/note="full consensus sequence for ribosome-binding sites
upstream of start codons in E. coli; complementary to a
region in the 3' end of the 16S rRNA (Chen et al., 1994)"
CDS 8898..8918
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
terminator 8957..9209
/label=NOS terminator
/note="nopaline synthase terminator and poly(A) signal"
primer_bind complement(9269..9285)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(9269..9285)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(9282..9304)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 9293..9309
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
promoter complement(9317..9347)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 9362..9383
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 9574..10250
/label=CaMV 35S promoter (enhanced)
/note="cauliflower mosaic virus 35S promoter with a
duplicated enhancer region"
CDS 10295..10846
/codon_start=1
/gene="Streptomyces hygroscopicus bar"
/product="phosphinothricin acetyltransferase"
/label=BlpR
/note="confers resistance to bialophos or phosphinothricin"
/translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE
WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS
TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF
WQLDFSLPVPPRPVLPVTEI"
polyA_signal 10853..11027
/label=CaMV poly(A) signal
/note="cauliflower mosaic virus polyadenylation signal"
misc_feature 11105..11129
/label=LB T-DNA repeat
/note="left border repeat from nopaline C58 T-DNA"
CDS 11554..12348
/codon_start=1
/gene="aphA-3"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin"
/translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
rep_origin 12435..13023
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 12924..12943
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 13177..13194
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatggt
61 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc
121 gctacgtgac tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg
181 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
241 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg aggcagggtg ccttgatgtg
301 ggcgccggcg gtcgagtggc gacggcgcgg cttgtccgcg ccctggtaga ttgcctggcc
361 gtaggccagc catttttgag cggccagcgg ccgcgatagg ccgacgcgaa gcggcggggc
421 gtagggagcg cagcgaccga agggtaggcg ctttttgcag ctcttcggct gtgcgctggc
481 cagacagtta tgcacaggcc aggcgggttt taagagtttt aataagtttt aaagagtttt
541 aggcggaaaa atcgcctttt ttctctttta tatcagtcac ttacatgtgt gaccggttcc
601 caatgtacgg ctttgggttc ccaatgtacg ggttccggtt cccaatgtac ggctttgggt
661 tcccaatgta cgtgctatcc acaggaaaga gaccttttcg acctttttcc cctgctaggg
721 caatttgccc tagcatctgc tccgtacatt aggaaccggc ggatgcttcg ccctcgatca
781 ggttgcggta gcgcatgact aggatcgggc cagcctgccc cgcctcctcc ttcaaatcgt
841 actccggcag gtcatttgac ccgatcagct tgcgcacggt gaaacagaac ttcttgaact
901 ctccggcgct gccactgcgt tcgtagatcg tcttgaacaa ccatctggct tctgccttgc
961 ctgcggcgcg gcgtgccagg cggtagagaa aacggccgat gccgggatcg atcaaaaagt
1021 aatcggggtg aaccgtcagc acgtccgggt tcttgccttc tgtgatctcg cggtacatcc
1081 aatcagctag ctcgatctcg atgtactccg gccgcccggt ttcgctcttt acgatcttgt
1141 agcggctaat caaggcttca ccctcggata ccgtcaccag gcggccgttc ttggccttct
1201 tcgtacgctg catggcaacg tgcgtggtgt ttaaccgaat gcaggtttct accaggtcgt
1261 ctttctgctt tccgccatcg gctcgccggc agaacttgag tacgtccgca acgtgtggac
1321 ggaacacgcg gccgggcttg tctcccttcc cttcccggta tcggttcatg gattcggtta
1381 gatgggaaac cgccatcagt accaggtcgt aatcccacac actggccatg ccggccggcc
1441 ctgcggaaac ctctacgtgc ccgtctggaa gctcgtagcg gatcacctcg ccagctcgtc
1501 ggtcacgctt cgacagacgg aaaacggcca cgtccatgat gctgcgacta tcgcgggtgc
1561 ccacgtcata gagcatcgga acgaaaaaat ctggttgctc gtcgcccttg ggcggcttcc
1621 taatcgacgg cgcaccggct gccggcggtt gccgggattc tttgcggatt cgatcagcgg
1681 ccgcttgcca cgattcaccg gggcgtgctt ctgcctcgat gcgttgccgc tgggcggcct
1741 gcgcggcctt caacttctcc accaggtcat cacccagcgc cgcgccgatt tgtaccgggc
1801 cggatggttt gcgaccgctc acgccgattc ctcgggcttg ggggttccag tgccattgca
1861 gggccggcag gcaacccagc cgcttacgcc tggccaaccg cccgttcctc cacacatggg
1921 gcattccacg gcgtcggtgc ctggttgttc ttgattttcc atgccgcctc ctttagccgc
1981 taaaattcat ctactcattt attcatttgc tcatttactc tggtagctgc gcgatgtatt
2041 cagatagcag ctcggtaatg gtcttgcctt ggcgtaccgc gtacatcttc agcttggtgt
2101 gatcctccgc cggcaactga aagttgaccc gcttcatggc tggcgtgtct gccaggctgg
2161 ccaacgttgc agccttgctg ctgcgtgcgc tcggacggcc ggcacttagc gtgtttgtgc
2221 ttttgctcat tttctcttta cctcattaac tcaaatgagt tttgatttaa tttcagcggc
2281 cagcgcctgg acctcgcggg cagcgtcgcc ctcgggttct gattcaagaa cggttgtgcc
2341 ggcggcggca gtgcctgggt agctcacgcg ctgcgtgata cgggactcaa gaatgggcag
2401 ctcgtacccg gccagcgcct cggcaacctc accgccgatg cgcgtgcctt tgatcgcccg
2461 cgacacgaca aaggccgctt gtagccttcc atccgtgacc tcaatgcgct gcttaaccag
2521 ctccaccagg tcggcggtgg cccatatgtc gtaagggctt ggctgcaccg gaatcagcac
2581 gaagtcggct gccttgatcg cggacacagc caagtccgcc gcctggggcg ctccgtcgat
2641 cactacgaag tcgcgccggc cgatggcctt cacgtcgcgg tcaatcgtcg ggcggtcgat
2701 gccgacaacg gttagcggtt gatcttcccg cacggccgcc caatcgcggg cactgccctg
2761 gggatcggaa tcgactaaca gaacatcggc cccggcgagt tgcagggcgc gggctagatg
2821 ggttgcgatg gtcgtcttgc ctgacccgcc tttctggtta agtacagcga taaccttcat
2881 gcgttcccct tgcgtatttg tttatttact catcgcatca tatacgcagc gaccgcatga
2941 cgcaagctgt tttactcaaa tacacatcac ctttttagac ggcggcgctc ggtttcttca
3001 gcggccaagc tggccggcca ggccgccagc ttggcatcag acaaaccggc caggatttca
3061 tgcagccgca cggttgagac gtgcgcgggc ggctcgaaca cgtacccggc cgcgatcatc
3121 tccgcctcga tctcttcggt aatgaaaaac ggttcgtcct ggccgtcctg gtgcggtttc
3181 atgcttgttc ctcttggcgt tcattctcgg cggccgccag ggcgtcggcc tcggtcaatg
3241 cgtcctcacg gaaggcaccg cgccgcctgg cctcggtggg cgtcacttcc tcgctgcgct
3301 caagtgcgcg gtacagggtc gagcgatgca cgccaagcag tgcagccgcc tctttcacgg
3361 tgcggccttc ctggtcgatc agctcgcggg cgtgcgcgat ctgtgccggg gtgagggtag
3421 ggcgggggcc aaacttcacg cctcgggcct tggcggcctc gcgcccgctc cgggtgcggt
3481 cgatgattag ggaacgctcg aactcggcaa tgccggcgaa cacggtcaac accatgcggc
3541 cggccggcgt ggtggtgtcg gcccacggct ctgccaggct acgcaggccc gcgccggcct
3601 cctggatgcg ctcggcaatg tccagtaggt cgcgggtgct gcgggccagg cggtctagcc
3661 tggtcactgt cacaacgtcg ccagggcgta ggtggtcaag catcctggcc agctccgggc
3721 ggtcgcgcct ggtgccggtg atcttctcgg aaaacagctt ggtgcagccg gccgcgtgca
3781 gttcggcccg ttggttggtc aagtcctggt cgtcggtgct gacgcgggca tagcccagca
3841 ggccagcggc ggcgctcttg ttcatggcgt aatgtctccg gttctagtcg caagtattct
3901 actttatgcg actaaaacac gcgacaagaa aacgccagga aaagggcagg gcggcagcct
3961 gtcgcgtaac ttaggacttg tgcgacatgt cgttttcaga agacggctgc actgaacgtc
4021 agaagccgac tgcactatag cagcggaggg gttggatcaa agtactttga tcccgagggg
4081 aaccctgtgg ttggcatgca catacaaatg gacgaacgga taaacctttt cacgcccttt
4141 taaatatccg attattctaa taaacgctct tttctcttag gtttacccgc caatatatcc
4201 tgtcaaacac tgatagttta aactgaaggc gggaaacgac aatctgatcc aagctcaagc
4261 tgctctagca ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct
4321 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa
4381 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgccaag cttgcatgcc
4441 tgcaggtcga ctctagagga tccccgggta ccgggccccc cctcgaggcg cgccaccagg
4501 tcgacatagt ttaaactgaa ggcgggaaac gacaatctga tccaagctca agctaagctt
4561 gcatgcctgc aggatatcgt ggatccaagc ttgccacgtg ccgccacgtg ccgccacgtg
4621 ccgccacgtg cctctagagg atccatctcc actgacgtaa gggatgacgc acaatcccac
4681 tatccttcgc aagacccttc ctctatataa ggaagttcat ttcatttgga gaggacacgc
4741 tgcttatgtc cagagctgat ccaaagaaga agagaaaggt ggtcgacatg gataagaagt
4801 actccatcgg actggatatc ggaactaact ccgtgggatg ggctgtgatc actgatgagt
4861 acaaggtgcc atccaagaag ttcaaggtgc tgggaaacac tgatagacac tccatcaaga
4921 agaacctgat cggagctctg ctgttcgatt ccggagagac tgctgaggct actagactga
4981 agagaactgc tagaagaaga tacactagaa gaaagaacag aatctgctac ctgcaagaga
5041 tcttctccaa cgagatggct aaggtggatg attccttctt ccacagactg gaggagtcct
5101 tcctggtgga ggaggataag aagcacgaga gacacccaat cttcggaaac atcgtggatg
5161 aggtggctta ccacgagaag tacccaacta tctaccacct gagaaagaag ctggtggatt
5221 ccactgataa ggctgatctg agactgatct acctggctct ggctcacatg atcaagttca
5281 gaggacactt cctgatcgag ggagatctga acccagataa ctccgatgtg gataagctgt
5341 tcatccaact ggtgcaaact tacaaccaac tgttcgagga gaacccaatc aacgcttccg
5401 gagtggatgc taaggctatc ctgtccgcta gactgtccaa gtccagaaga ctggagaacc
5461 tgatcgctca actgccagga gagaagaaga acggactgtt cggaaacctg atcgctctgt
5521 ccctgggact gactccaaac ttcaagtcca acttcgatct ggctgaggat gctaagctgc
5581 aactgtccaa ggatacttac gatgatgatc tggataacct gctggctcaa atcggagatc
5641 aatacgctga tctgttcctg gctgctaaga acctgtccga tgctatcctg ctgtccgata
5701 tcctgagagt gaacactgag atcactaagg ctccactgtc cgcttccatg atcaagagat
5761 acgatgagca ccaccaagat ctgactctgc tgaaggctct ggtgagacaa caactgccag
5821 agaagtacaa ggagatcttc ttcgatcaat ccaagaacgg atacgctgga tacatcgatg
5881 gaggagcttc ccaagaggag ttctacaagt tcatcaagcc aatcctggag aagatggatg
5941 gaactgagga gctgctggtg aagctgaaca gagaggatct gctgagaaag caaagaactt
6001 tcgataacgg atccatccca caccaaatcc acctgggaga gctgcacgct atcctgagaa
6061 gacaagagga tttctaccca ttcctgaagg ataacagaga gaagatcgag aagatcctga
6121 ctttcagaat cccatactac gtgggaccac tggctagagg aaactccaga ttcgcttgga
6181 tgactagaaa gtccgaggag actatcactc catggaactt cgaggaggtg gtggataagg
6241 gagcttccgc tcaatccttc atcgagagaa tgactaactt cgataagaac ctgccaaacg
6301 agaaggtgct gccaaagcac tccctgctgt acgagtactt cactgtgtac aacgagctga
6361 ctaaggtgaa gtacgtgact gagggaatga gaaagccagc tttcctgtcc ggagagcaaa
6421 agaaggctat cgtggatctg ctgttcaaga ctaacagaaa ggtgactgtg aagcaactga
6481 aggaggatta cttcaagaag atcgagtgct tcgattccgt ggagatctcc ggagtggagg
6541 atagattcaa cgcttccctg ggaacttacc acgatctgct gaagatcatc aaggataagg
6601 atttcctgga taacgaggag aacgaggata tcctggagga tatcgtgctg actctgactc
6661 tgttcgagga tagagagatg atcgaggaga gactgaagac ttacgctcac ctgttcgatg
6721 ataaggtgat gaagcaactg aagagaagaa gatacactgg atggggaaga ctgtccagaa
6781 agctgatcaa cggaatcaga gataagcaat ccggaaagac tatcctggat ttcctgaagt
6841 ccgatggatt cgctaacaga aacttcatgc aactgatcca cgatgattcc ctgactttca
6901 aggaggatat ccaaaaggct caagtgtccg gacaaggaga ttccctgcac gagcacatcg
6961 ctaacctggc tggatcccca gctatcaaga agggaatcct gcaaactgtg aaggtggtgg
7021 atgagctggt gaaggtgatg ggaagacaca agccagagaa catcgtgatc gagatggcta
7081 gagagaacca aactactcaa aagggacaaa agaactccag agagagaatg aagagaatcg
7141 aggagggaat caaggagctg ggatcccaaa tcctgaagga gcacccagtg gagaacactc
7201 aactgcaaaa cgagaagctg tacctgtact acctgcaaaa cggaagagat atgtacgtgg
7261 atcaagagct ggatatcaac agactgtccg attacgatgt ggatcacatc gtgccacaat
7321 ccttcctgaa ggatgattcc atcgataaca aggtgctgac tagatccgat aagaacagag
7381 gaaagtccga taacgtgcca tccgaggagg tggtgaagaa gatgaagaac tactggagac
7441 aactgctgaa cgctaagctg atcactcaaa gaaagttcga taacctgact aaggctgaga
7501 gaggaggact gtccgagctg gataaggctg gattcatcaa gagacaactg gtggagacta
7561 gacaaatcac taagcacgtg gctcaaatcc tggattccag aatgaacact aagtacgatg
7621 agaacgataa gctgatcaga gaggtgaagg tgatcactct gaagtccaag ctggtgtccg
7681 atttcagaaa ggatttccaa ttctacaagg tgagagagat caacaactac caccacgctc
7741 acgatgctta cctgaacgct gtggtgggaa ctgctctgat caagaagtac ccaaagctgg
7801 agtccgagtt cgtgtacgga gattacaagg tgtacgatgt gagaaagatg atcgctaagt
7861 ccgagcaaga gatcggaaag gctactgcta agtacttctt ctactccaac atcatgaact
7921 tcttcaagac tgagatcact ctggctaacg gagagatcag aaagagacca ctgatcgaga
7981 ctaacggaga gactggagag atcgtgtggg ataagggaag agatttcgct actgtgagaa
8041 aggtgctgtc catgccacaa gtgaacatcg tgaagaagac tgaggtgcaa actggaggat
8101 tctccaagga gtccatcctg ccaaagagaa actccgataa gctgatcgct agaaagaagg
8161 attgggatcc aaagaagtac ggaggattcg attccccaac tgtggcttac tccgtgctgg
8221 tggtggctaa ggtggagaag ggaaagtcca agaagctgaa gtccgtgaag gagctgctgg
8281 gaatcactat catggagaga tcctccttcg agaagaaccc aatcgatttc ctggaggcta
8341 agggatacaa ggaggtgaag aaggatctga tcatcaagct gccaaagtac tccctgttcg
8401 agctggagaa cggaagaaag agaatgctgg cttccgctgg agagctgcaa aagggaaacg
8461 agctggctct gccatccaag tacgtgaact tcctgtacct ggcttcccac tacgagaagc
8521 tgaagggatc cccagaggat aacgagcaaa agcaactgtt cgtggagcaa cacaagcact
8581 acctggatga gatcatcgag caaatctccg agttctccaa gagagtgatc ctggctgatg
8641 ctaacctgga taaggtgctg tccgcttaca acaagcacag agataagcca atcagagagc
8701 aagctgagaa catcatccac ctgttcactc tgactaacct gggagctcca gctgctttca
8761 agtacttcga tactactatc gatagaaaga gatacacttc cactaaggag gtgctggatg
8821 ctactctgat ccaccaatcc atcactggac tgtacgagac tagaatcgat ctgtcccaac
8881 tgggaggaga tctgcaacca aagaagaaga gaaaggtggg aggatgatct agaccagatt
8941 aattaagaat ttccccgatc gttcaaacat ttggcaataa agtttcttaa gattgaatcc
9001 tgttgccggt cttgcgatga ttatcatata atttctgttg aattacgtta agcatgtaat
9061 aattaacatg taatgcatga cgttatttat gagatgggtt tttatgatta gagtcccgca
9121 attatacatt taatacgcga tagaaaacaa aatatagcgc gcaaactagg ataaattatc
9181 gcgcgcggtg tcatctatgt tactagatcg gttaattaac tagttctaga gcggccgcca
9241 ccgcggtgga gctcgaattc gtaatcatgt catagctgtt tcctgtgtga aattgttatc
9301 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
9361 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
9421 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
9481 ttggctagag cagcttgcca acatggtgga gcacgacact ctcgtctact ccaagaatat
9541 caaagataca gtctcagaag accaaagggc tattgagact tttcaacaaa gggtaatatc
9601 gggaaacctc ctcggattcc attgcccagc tatctgtcac ttcatcaaaa ggacagtaga
9661 aaaggaaggt ggcacctaca aatgccatca ttgcgataaa ggaaaggcta tcgttcaaga
9721 tgcctctgcc gacagtggtc ccaaagatgg acccccaccc acgaggagca tcgtggaaaa
9781 agaagacgtt ccaaccacgt cttcaaagca agtggattga tgtgaacatg gtggagcacg
9841 acactctcgt ctactccaag aatatcaaag atacagtctc agaagaccaa agggctattg
9901 agacttttca acaaagggta atatcgggaa acctcctcgg attccattgc ccagctatct
9961 gtcacttcat caaaaggaca gtagaaaagg aaggtggcac ctacaaatgc catcattgcg
10021 ataaaggaaa ggctatcgtt caagatgcct ctgccgacag tggtcccaaa gatggacccc
10081 cacccacgag gagcatcgtg gaaaaagaag acgttccaac cacgtcttca aagcaagtgg
10141 attgatgtga tatctccact gacgtaaggg atgacgcaca atcccactat ccttcgcaag
10201 acccttcctc tatataagga agttcatttc atttggagag gacacgctga aatcaccagt
10261 ctctctctac aaatctatct ctctcgagtc taccatgagc ccagaacgac gcccggccga
10321 catccgccgt gccaccgagg cggacatgcc ggcggtctgc accatcgtca accactacat
10381 cgagacaagc acggtcaact tccgtaccga gccgcaggaa ccgcaggagt ggacggacga
10441 cctcgtccgt ctgcgggagc gctatccctg gctcgtcgcc gaggtggacg gcgaggtcgc
10501 cggcatcgcc tacgcgggcc cctggaaggc acgcaacgcc tacgactgga cggccgagtc
10561 gaccgtgtac gtctcccccc gccaccagcg gacgggactg ggctccacgc tctacaccca
10621 cctgctgaag tccctggagg cacagggctt caagagcgtg gtcgctgtca tcgggctgcc
10681 caacgacccg agcgtgcgca tgcacgaggc gctcggatat gccccccgcg gcatgctgcg
10741 ggcggccggc ttcaagcacg ggaactggca tgacgtgggt ttctggcagc tggacttcag
10801 cctgccggta ccgccccgtc cggtcctgcc cgtcaccgag atttgactcg agtttctcca
10861 taataatgtg tgagtagttc ccagataagg gaattagggt tcctataggg tttcgctcat
10921 gtgttgagca tataagaaac ccttagtatg tatttgtatt tgtaaaatac ttctatcaat
10981 aaaatttcta attcctaaaa ccaaaatcca gtactaaaat ccagatcccc cgaattaatt
11041 cggcgttaat tcagtacatt aaaaacgtcc gcaatgtgtt attaagttgt ctaagcgtca
11101 atttgtttac accacaatat atcctgccac cagccagcca acagctcccc gaccggcagc
11161 tcggcacaaa atcaccactc gatacaggca gcccatcagt ccgggacggc gtcagcggga
11221 gagccgttgt aaggcggcag actttgctca tgttaccgat gctattcgga agaacggcaa
11281 ctaagctgcc gggtttgaaa cacggatgat ctcgcggagg gtagcatgtt gattgtaacg
11341 atgacagagc gttgctgcct gtgatcaccg cggtttcaaa atcggctccg tcgatactat
11401 gttatacgcc aactttgaaa acaactttga aaaagctgtt ttctggtatt taaggtttta
11461 gaatgcaagg aacagtgaat tggagttcgt cttgttataa ttagcttctt ggggtatctt
11521 taaatactgt agaaaagagg aaggaaataa taaatggcta aaatgagaat atcaccggaa
11581 ttgaaaaaac tgatcgaaaa ataccgctgc gtaaaagata cggaaggaat gtctcctgct
11641 aaggtatata agctggtggg agaaaatgaa aacctatatt taaaaatgac ggacagccgg
11701 tataaaggga ccacctatga tgtggaacgg gaaaaggaca tgatgctatg gctggaagga
11761 aagctgcctg ttccaaaggt cctgcacttt gaacggcatg atggctggag caatctgctc
11821 atgagtgagg ccgatggcgt cctttgctcg gaagagtatg aagatgaaca aagccctgaa
11881 aagattatcg agctgtatgc ggagtgcatc aggctctttc actccatcga catatcggat
11941 tgtccctata cgaatagctt agacagccgc ttagccgaat tggattactt actgaataac
12001 gatctggccg atgtggattg cgaaaactgg gaagaagaca ctccatttaa agatccgcgc
12061 gagctgtatg attttttaaa gacggaaaag cccgaagagg aacttgtctt ttcccacggc
12121 gacctgggag acagcaacat ctttgtgaaa gatggcaaag taagtggctt tattgatctt
12181 gggagaagcg gcagggcgga caagtggtat gacattgcct tctgcgtccg gtcgatcagg
12241 gaggatatcg gggaagaaca gtatgtcgag ctattttttg acttactggg gatcaagcct
12301 gattgggaga aaataaaata ttatatttta ctggatgaat tgttttagta cctagaatgc
12361 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
12421 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
12481 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
12541 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
12601 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
12661 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
12721 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
12781 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
12841 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
12901 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
12961 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
13021 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
13081 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
13141 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
13201 gaag
//
