| 质粒名称: | p-dCas9-SSAP-MS2-BB_BbsI |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 183826 |
| 存储实验室: | Le Cong |
|
载体骨架基本信息 |
|
| 载体名称: | pCBH |
| 载体出品公司: | -- |
| 空载体大小: | -- |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | CRISPR |
| 筛选标记: | -- |
| 载体抗性: | 氨苄霉素100 μg/mL |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | U6-guideRNA-dCas9-T2A-EBFP |
| 物种来源: | Synthetic |
| 插入大小: | 6291 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子: | CBH |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | NcoI（如果已销毁，则未知） |
| 3’克隆位点: | EcoRI（如果已销毁，则未知） |
| 5’测序引物 | pLKO |
| 3’测序引物 | BHGpA |
|
文献引用方法 |
|
| 材料和方法部分: | p-dCas9-SSAP-MS2-BB_BbsI was a gift from Le Cong (Addgene plasmid # 183826 ; http://n2t.net/addgene:183826 ; RRID:Addgene_183826) |
| 参考文献部分: | dCas9-based gene editing for cleavage-free genomic knock-in of long sequences. Wang C, Qu Y, Cheng JKW, Hughes NW, Zhang Q, Wang M, Cong L. Nat Cell Biol. 2022 Feb;24(2):268-278. doi: 10.1038/s41556-021-00836-1. Epub 2022 Feb 10. 10.1038/s41556-021-00836-1 PubMed 35145221 |
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 p-dCas9-SSAP-MS2-BB_BbsI 9277 bp ds-DNA circular SYN 05-MAY-2022
DEFINITION pU6-MS2-gRNA-backbone(BbsI)-CBH-dSpCas9-T2A-EBFP.
ACCESSION .
VERSION .
KEYWORDS p-dCas9-SSAP-MS2-BB_BbsI
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9277)
AUTHORS Wang C, Qu Y, Cheng JKW, Hughes NW, Zhang Q, Wang M, Cong L
TITLE dCas9-based gene editing for cleavage-free genomic knock-in of long
sequences.
JOURNAL Nat Cell Biol. 2022 Feb;24(2):268-278. doi:
10.1038/s41556-021-00836-1. Epub 2022 Feb 10.
PUBMED 35145221
REFERENCE 2 (bases 1 to 9277)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 5, 2022 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..9277
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..241
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 1..21
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 172..191
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_RNA 285..303
/label=MS2 stem loop
/note="stem loop that binds the bacteriophage MS2 coat
protein"
misc_RNA 378..396
/label=MS2 stem loop
/note="stem loop that binds the bacteriophage MS2 coat
protein"
enhancer 417..702
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer;
contains an 18-bp deletion relative to the standard CMV
enhancer"
promoter 704..981
/label=chicken beta-actin promoter
intron 982..1209
/label=hybrid intron
/note="hybrid between chicken beta-actin (CBA) and minute
virus of mice (MMV) introns (Gray et al., 2011)"
regulatory 1221..1230
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 1230..1295
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
CDS 1302..1322
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 1347..5447
/codon_start=1
/product="catalytically dead mutant of the Cas9
endonuclease from the Streptococcus pyogenes Type II
CRISPR/Cas system"
/label=dCas9
/note="RNA-guided DNA-binding protein that lacks
endonuclease activity due to the D10A mutation in the RuvC
catalytic domain and the H840A mutation in the HNH
catalytic domain"
/translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 5448..5495
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
CDS 5511..5564
/codon_start=1
/product="2A peptide from Thosea asigna virus capsid
protein"
/label=T2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="EGRGSLLTCGDVEENPGP"
CDS 5571..6290
/codon_start=1
/product="enhanced blue variant of GFP (Ai et al., 2007)"
/label=EBFP2
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK
VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL
EFRTAAGITLGMDELYK"
primer_bind complement(5616..5637)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(6318..6335)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 6324..6531
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
repeat_region 6540..6680
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 6540..6669
/label=AAV2 ITR
rep_origin 6755..7210
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(6842..6861)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 7052..7073
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(7227..7246)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 7346..7368
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(7406..7424)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 7492..7596
/gene="bla"
/label=AmpR promoter
CDS 7597..8457
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(7815..7834)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 8628..9216
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 9117..9136
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtggcgtaga
121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
241 cgaaacaccg ggtcttcgag aagacctgtt taagagctag gccaacatga ggatcaccca
301 tgtctgcagg gcctagcaag tttaaataag gctagtccgt tatcaacttg aaaaagtggc
361 accgagtcgg tgcgcgcaca tgaggatcac ccatgtgctt tttttctaga ggtacccgtt
421 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
481 tcaatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
541 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
601 aatgacggta aatggcccgc ctggcattgt gcccagtaca tgaccttatg ggactttcct
661 acttggcagt acatctacgt attagtcatc gctattacca tggtcgaggt gagccccacg
721 ttctgcttca ctctccccat ctcccccccc tccccacccc caattttgta tttatttatt
781 ttttaattat tttgtgcagc gatgggggcg gggggggggg gggggcgcgc gccrggsggg
841 gsggggsggg gsgrggggsg gggsggggcg aggcggagag gtgcggcggc agccaatcag
901 agcggcgcgc tccgaaagtt tccttttatg gcgaggcggc ggcggcggcg gccctataaa
961 aagcgaagcg cgcggcgggc gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc
1021 gccgccgcct cgcgccgccc gccccggctc tgactgaccg cgttactccc acaggtgagc
1081 gggcgggacg gcccttctcc tccgggctgt aattagctga gcaagaggta agggtttaag
1141 ggatggttgg ttggtggggt attaatgttt aattacctgg agcacctgcc tgaaatcact
1201 ttttttcagg ttggaccggt gccaccatgg actataagga ccacgacgga gactacaagg
1261 atcatgatat tgattacaaa gacgatgacg ataagatggc cccaaagaag aagcggaagg
1321 tcggtatcca cggagtccca gcagccgaca agaagtacag catcggcctg gccatcggca
1381 ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca
1441 aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt
1501 tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca
1561 ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg
1621 tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc
1681 acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc
1741 ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc
1801 tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg
1861 acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca
1921 accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt
1981 ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga
2041 agaagaatgg cctgttcggc aacctgattg ccctgagcct gggcctgacc cccaacttca
2101 agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg
2161 acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg
2221 ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca
2281 ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga
2341 ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg
2401 accagagcaa gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct
2461 acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc
2521 tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc
2581 agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc
2641 tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg
2701 gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca
2761 tcaccccctg gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg
2821 agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc
2881 tgctgtacga gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg
2941 gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt
3001 tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg
3061 agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc tccctgggca
3121 cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat gaggaaaacg
3181 aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga gagatgatcg
3241 aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag cagctgaagc
3301 ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc atccgggaca
3361 agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc aacagaaact
3421 tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag aaagcccagg
3481 tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc agccccgcca
3541 ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc
3601 ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc acccagaagg
3661 gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa gagctgggca
3721 gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag aagctgtacc
3781 tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac atcaaccggc
3841 tgtccgacta cgatgtggac gccatcgtgc ctcagagctt tctgaaggac gactccatcg
3901 acaacaaggt gctgaccaga agcgacaaga accggggcaa gagcgacaac gtgccctccg
3961 aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc aagctgatta
4021 cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc gaactggata
4081 aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag cacgtggcac
4141 agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg atccgggaag
4201 tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat ttccagtttt
4261 acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg aacgccgtcg
4321 tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg tacggcgact
4381 acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta
4441 ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag attaccctgg
4501 ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc ggggagatcg
4561 tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg ccccaagtga
4621 atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct atcctgccca
4681 agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag aagtacggcg
4741 gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg gaaaagggca
4801 agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg gaaagaagca
4861 gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg
4921 acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc cggaagagaa
4981 tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc tccaaatatg
5041 tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc gaggataatg
5101 agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc atcgagcaga
5161 tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg
5221 cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc atccacctgt
5281 ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc accatcgacc
5341 ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac cagagcatca
5401 ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacaaa aggccggcgg
5461 ccacgaaaaa ggccggccag gcaaaaaaga aaaaggaatt cggcagtgga gagggcagag
5521 gaagtctgct aacatgcggt gacgtggagg agaatcccgg ccctgctagc atggtgagca
5581 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
5641 acggccacaa gttcagcgtg aggggcgagg gcgagggcga tgccaccaac ggcaagctga
5701 ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
5761 ccctgagcca cggcgtgcag tgcttcgccc gctaccccga ccacatgaag cagcacgact
5821 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
5881 acggcaccta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
5941 tcgagctgaa gggcgtcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
6001 acaacttcaa cagccacaac atctatatca tggccgtcaa gcagaagaac ggcatcaagg
6061 tgaacttcaa gatccgccac aacgtggagg acggcagcgt gcagctcgcc gaccactacc
6121 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacagccac tacctgagca
6181 cccagtccgt gctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
6241 tccgcaccgc cgccgggatc actctcggca tggacgagct gtacaagtga gaattctaac
6301 tagagctcgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
6361 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
6421 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
6481 gcaggacagc aagggggagg attgggaaga gaatagcagg catgctgggg agcggccgca
6541 ggaaccccta gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc
6601 cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg
6661 agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg
6721 tatttcacac cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc
6781 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cttagcgccc
6841 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
6901 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
6961 aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
7021 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
7081 ctcaactcta tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat
7141 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
7201 tttacaattt tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
7261 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
7321 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
7381 tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc
7441 atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
7501 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
7561 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
7621 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
7681 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
7741 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
7801 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
7861 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
7921 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
7981 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
8041 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
8101 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
8161 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
8221 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
8281 attgctgata aatctggagc cggtgagcgt ggaagccgcg gtatcattgc agcactgggg
8341 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
8401 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
8461 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
8521 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
8581 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
8641 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
8701 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
8761 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
8821 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
8881 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
8941 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
9001 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
9061 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
9121 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
9181 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
9241 cggttcctgg ccttttgctg gccttttgct cacatgt
//
