| 质粒名称: | AAVS1-Blasticidin-CAG-Flpe-ERT2 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 68461 |
| 存储实验室: | Su-Chun Zhang |
|
载体骨架基本信息 |
|
| 载体名称: | AAVS1-CAG |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 8085 bp |
| 完整质粒大小: | 12382 bp |
| 载体修饰: | amplified mammalian-codon-optimized Flpe recombinase cDNA and ERT2 cDNA by PCR from pDIRE (Addgene plasmid #26745) and pCAG-FlpeERT2 (Addgene plasmid #14756), respectively. The mammalian codon-optimized Flpe fused with ERT2 was inserted into the AAVS1-neo-CAG-hrGFP to replace GFP to get the AAVS1-neo-CAG-FlpeERT2 donor plasmid. |
| 载体类型: | 哺乳细胞表达,合成生物学 |
| 筛选标记: | 杀稻瘟菌素 |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | mammalian-codon-optimized Flpe recombinase fused with ERT2 |
| 物种来源: | Synthetic |
| 插入大小: | 2243 bp |
| 启动子 | CAG |
| 标签/融合蛋白: | ERT2（插入件上的C端子） |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | SalI (未破坏) |
| 3' 克隆位点: | MluI (未破坏) |
| 5' 测序引物 | ggcttctggcgtgtgaccggc |
| 3' 测序引物 | catagcgtaaaaggagcaaca |
|
文献引用方法 |
|
| 材料和方法部分: | AAVS1-Blasticidin-CAG-Flpe-ERT2 was a gift from Su-Chun Zhang (Addgene plasmid # 68461 ; http://n2t.net/addgene:68461 ; RRID:Addgene_68461) |
| 参考文献部分: | Engineering Human Stem Cell Lines with Inducible Gene Knockout using CRISPR/Cas9. Chen Y, Cao J, Xiong M, Petersen AJ, Dong Y, Tao Y, Huang CT, Du Z, Zhang SC. Cell Stem Cell. 2015 Jul 1. pii: S1934-5909(15)00261-1. doi: 10.1016/j.stem.2015.06.001. 10.1016/j.stem.2015.06.001 PubMed 26145478 |
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 AAVS1-Blasticidin-CAG-Flpe-ERT2 12382 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION AAVS1 targeting donor plasmid with Flpe-ERT2 expression cassette and
blasticidin selection gene.
ACCESSION .
VERSION .
KEYWORDS AAVS1-Blasticidin-CAG-Flpe-ERT2
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 12382)
AUTHORS Chen Y, Cao J, Xiong M, Petersen AJ, Dong Y, Tao Y, Huang CT, Du Z,
Zhang SC
TITLE Engineering Human Stem Cell Lines with Inducible Gene Knockout using
CRISPR/Cas9.
JOURNAL Cell Stem Cell. 2015 Jul 1. pii: S1934-5909(15)00261-1. doi:
10.1016/j.stem.2015.06.001.
PUBMED 26145478
REFERENCE 2 (bases 1 to 12382)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..12382
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_feature 33..836
/label=HA-L
/note="left homology arm from the adeno-associated virus
integration site (AAVS1) within intron 1 of the human
PPP1R12C gene"
misc_feature 843..868
/label=SA
/note="splice acceptor site"
CDS 892..945
/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"
primer_bind complement(1418..1435)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 1424..1648
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
enhancer 1716..2095
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 2097..2374
/label=chicken beta-actin promoter
intron 2376..3393
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
primer_bind 3317..3339
/label=pCAGGS-5
/note="Chimeric intron in CAG promoter, forward primer"
primer_bind 3401..3420
/label=pCAG-F
/note="Rabbit beta-globin intron, for pCAG plasmids,
forward primer"
regulatory 3457..3466
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 4762..5697
/codon_start=1
/product="mutated ligand-binding domain of the human
estrogen receptor (Feil et al., 1997)"
/label=ERT2
/translation="AGDMRAANLWPSPLMIKRSKKNSLALSLTADQMVSALLDAEPPIL
YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEI
LMIGLVWRSMEHPVKLLFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEF
VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL
AQLLLILSHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEAADAHRLHAPTSRGGASVEET
DQSHLATAGSTSSHSLQKYYITGEAEGFPAT"
misc_feature 5737..6325
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(5790..5810)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(6208..6219)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
polyA_signal 6357..6833
/label=hGH poly(A) signal
/note="human growth hormone polyadenylation signal"
primer_bind complement(6556..6575)
/label=hGH-PA-R
/note="Human growth hormone terminator, reverse primer"
primer_bind complement(6943..6962)
/label=Bglob-pA-R
/note="Rabbit beta-globin polyA region, reverse primer"
polyA_site 7008..7063
/label=-beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal"
primer_bind complement(7062..7081)
/label=rbglobpA-R
/note="Rabbit beta-globin polyA, reverse primer. Also
called rb-glob-pA-term-R"
primer_bind complement(7424..7440)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 7448..7464
/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(7472..7502)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 7517..7538
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
misc_feature 7612..8448
/label=HA-R
/note="right homology arm from the adeno-associated virus
integration site (AAVS1) within intron 1 of the human
PPP1R12C gene"
primer_bind complement(8488..8507)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(8489..8507)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(8514..8531)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(8514..8530)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(8523..8545)
/label=M13/pUC Forward
/note="In lacZ gene"
CDS 8668..8970
/codon_start=1
/gene="ccdB"
/product="CcdB, a bacterial toxin that poisons DNA gyrase"
/label=ccdB
/note="Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains."
/translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
primer_bind 8849..8868
/label=ccdB-fwd
/note="ccdB gene, forward primer"
primer_bind complement(9373..9392)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 9987..10006
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
CDS complement(10367..11227)
/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 10990..11009
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 11351..11939
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 11840..11859
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 12093..12110
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 12227..12248
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 12263..12293
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 12301..12317
/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)."
primer_bind 12325..12341
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
promoter 12362..12380
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
1 ctagtcctgc aggtttaaac gaattcgccc tttgctttct ctgaccagca ttctctcccc
61 tgggcctgtg ccgctttctg tctgcagctt gtggcctggg tcacctctac ggctggccca
121 gatccttccc tgccgcctcc ttcaggttcc gtcttcctcc actccctctt ccccttgctc
181 tctgctgtgt tgctgcccaa ggatgctctt tccggagcac ttccttctcg gcgctgcacc
241 acgtgatgtc ctctgagcgg atcctccccg tgtctgggtc ctctccgggc atctctcctc
301 cctcacccaa ccccatgccg tcttcactcg ctgggttccc ttttccttct ccttctgggg
361 cctgtgccat ctctcgtttc ttaggatggc cttctccgac ggatgtctcc cttgcgtccc
421 gcctcccctt cttgtaggcc tgcatcatca ccgtttttct ggacaacccc aaagtacccc
481 gtctccctgg ctttagccac ctctccatcc tcttgctttc tttgcctgga caccccgttc
541 tcctgtggat tcgggtcacc tctcactcct ttcatttggg cagctcccct acccccctta
601 cctctctagt ctgtgctagc tcttccagcc ccctgtcatg gcatcttcca ggggtccgag
661 agctcagcta gtcttcttcc tccaacccgg gcccctatgt ccacttcagg acagcatgtt
721 tgctgcctcc agggatcctg tgtccccgag ctgggaccac cttatattcc cagggccggt
781 taatgtggct ctggttctgg gtacttttat ctgtcccctc caccccacag tggggcaagc
841 ttctgacctc ttctcttcct cccacagggc ctcgagagat ctggcagcgg agagggcaga
901 ggaagtcttc taacatgcgg tgacgtggag gagaatcccg gccctaggac cggtatgaaa
961 acatttaaca tttctcaaca agatctagaa ttagtagaag tagcgacaga gaagattaca
1021 atgctttatg aggataataa acatcatgtg ggagcggcaa ttcgtacgaa aacaggagaa
1081 atcatttcgg cagtacatat tgaagcgtat ataggacgag taactgtttg tgcagaagcc
1141 attgcgattg gtagtgcagt ttcgaatgga caaaaggatt ttgacacgat tgtagctgtt
1201 agacaccctt attctgacga agtagataga agtattcgag tggtaagtcc ttgtggtatg
1261 tgtagggagt tgatttcaga ctatgcacca gattgttttg tgttaataga aatgaatggc
1321 aagttagtca aaactacgat tgaagaactc attccactca aatatacccg aaattaatga
1381 gaattcgggg atcaattctc tagagctcgc tgatcagcct cgactgtgcc ttctagttgc
1441 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
1501 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
1561 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
1621 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctcg
1681 actagagctt gcggaaccct taatggtacc gtctcgacat tgattattga ctagttatta
1741 atagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata
1801 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat
1861 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga
1921 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc
1981 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
2041 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtcga
2101 ggtgagcccc acgttctgct tcactctccc catctccccc ccctccccac ccccaatttt
2161 gtatttattt attttttaat tattttgtgc agcgatgggg gcgggggggg ggggggggcg
2221 cgcgccaggc ggggcggggc ggggcgaggg gcggggcggg gcgaggcgga gaggtgcggc
2281 ggcagccaat cagagcggcg cgctccgaaa gtttcctttt atggcgaggc ggcggcggcg
2341 gcggccctat aaaaagcgaa gcgcgcggcg ggcggggagt cgctgcgacg ctgccttcgc
2401 cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta
2461 ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt
2521 taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct ccgggagggc
2581 cctttgtgcg gggggagcgg ctcggggggt gcgtgcgtgt gtgtgtgcgt ggggagcgcc
2641 gcgtgcggct ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc ggggctttgt
2701 gcgctccgca gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg gtgcgggggg
2761 ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg tgggggggtg agcagggggt
2821 gtgggcgcgt cggtcgggct gcaacccccc ctgcaccccc ctccccgagt tgctgagcac
2881 ggcccggctt cgggtgcggg gctccgtacg gggcgtggcg cggggctcgc cgtgccgggc
2941 ggggggtggc ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc cggggagggc
3001 tcgggggagg ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc ggcgagccgc
3061 agccattgcc ttttatggta atcgtgcgag agggcgcagg gacttccttt gtcccaaatc
3121 tgtgcggagc cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg cggggcgaag
3181 cggtgcggcg ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc gccgcgccgc
3241 cgtccccttc tccctctcca gcctcggggc tgtccgcggg gggacggctg ccttcggggg
3301 ggacggggca gggcggggtt cggcttctgg cgtgtgaccg gcggctctag agcctctgct
3361 aaccatgttc atgccttctt ctttttccta cagctcctgg gcaacgtgct ggttattgtg
3421 ctgtctcatc attttggcaa agaattcgtc gacgccgcca ccatggctat gagccagttc
3481 gacatcctgt gcaagacccc ccccaaggtg ctggtgcggc agttcgtgga gagattcgag
3541 aggcccagcg gcgagaagat cgccagctgt gccgccgagc tgacctacct gtgctggatg
3601 atcacccaca acggcaccgc catcaagagg gccaccttca tgagctacaa caccatcatc
3661 agcaacagcc tgagcttcga catcgtgaac aagagcctgc agttcaagta caagacccag
3721 aaggccacca tcctggaggc cagcctgaag aagctgatcc ccgcctggga gttcaccatc
3781 atcccttaca acggccagaa gcaccagagc gacatcaccg acatcgtgtc cagcctgcag
3841 ctgcagttcg agagcagcga ggaggccgac aagggcaaca gccacagcaa gaagatgctg
3901 aaggccctgc tgtccgaggg cgagagcatc tgggagatca ccgagaagat cctgaacagc
3961 ttcgagtaca ccagcaggtt caccaagacc aagaccctgt accagttcct gttcctggcc
4021 acattcatca actgcggcag gttcagcgac atcaagaacg tggaccccaa gagcttcaag
4081 ctggtgcaga acaagtacct gggcgtgatc attcagtgcc tggtgaccga gaccaagaca
4141 agcgtgtcca ggcacatcta ctttttcagc gccagaggca ggatcgaccc cctggtgtac
4201 ctggacgagt tcctgaggaa cagcgagccc gtgctgaaga gagtgaacag gaccggcaac
4261 agcagcagca acaagcagga gtaccagctg ctgaaggaca acctggtgcg cagctacaac
4321 aaggccctga agaagaacgc cccctacccc atcttcgcta tcaagaacgg ccctaagagc
4381 cacatcggca ggcacctgat gaccagcttt ctgagcatga agggcctgac cgagctgaca
4441 aacgtggtgg gcaactggag cgacaagagg gcctccgccg tggccaggac cacctacacc
4501 caccagatca ccgccatccc cgaccactac ttcgccctgg tgtccaggta ctacgcctac
4561 gaccccatca gcaaggagat gatcgccctg aaggacgaga ccaaccccat cgaggagtgg
4621 cagcacatcg agcagctgaa gggcagcgcc gagggcagca tcagataccc cgcctggaac
4681 ggcatcatca gccaggaggt gctggactac ctgagcagct acatcaacag gcggatcacc
4741 ggttgcgtac gcggatcctc tgctggagac atgagagctg ccaacctttg gccaagcccg
4801 ctcatgatca aacgctctaa gaagaacagc ctggccttgt ccctgacggc cgaccagatg
4861 gtcagtgcct tgttggatgc tgagcccccc atactctatt ccgagtatga tcctaccaga
4921 cccttcagtg aagcttcgat gatgggctta ctgaccaacc tggcagacag ggagctggtt
4981 cacatgatca actgggcgaa gagggtgcca ggctttgtgg atttgaccct ccatgatcag
5041 gtccaccttc tagaatgtgc ctggctagag atcctgatga ttggtctcgt ctggcgctcc
5101 atggagcacc cagtgaagct actgtttgct cctaacttgc tcttggacag gaaccaggga
5161 aaatgtgtag agggcatggt ggagatcttc gacatgctgc tggctacatc atctcggttc
5221 cgcatgatga atctgcaggg agaggagttt gtgtgcctca aatctattat tttgcttaat
5281 tctggagtgt acacatttct gtccagcacc ctgaagtctc tggaagagaa ggaccatatc
5341 caccgagtcc tggacaagat cacagacact ttgatccacc tgatggccaa ggcaggcctg
5401 accctgcagc agcagcacca gcggctggcc cagctcctcc tcatcctctc ccacatcagg
5461 cacatgagta acaaaggcat ggagcatctg tacagcatga agtgcaagaa cgtggtgccc
5521 ctctatgacc tgctgctgga ggcggcggac gcccaccgcc tacatgcgcc cactagccgt
5581 ggaggggcat ccgtggagga gacggaccaa agccacttgg ccactgcggg ctctacttca
5641 tcgcattcct tgcaaaagta ttacatcacg ggggaggcag agggtttccc tgccacagct
5701 tgatagacgc gtgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt
5761 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
5821 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
5881 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
5941 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
6001 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
6061 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
6121 ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg
6181 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
6241 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
6301 atctcccttt gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt
6361 ggcatccctg tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc
6421 ccaccagcct tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct
6481 ataatattat ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct
6541 gtagggcctg cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca
6601 ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg
6661 gattccaggc atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt
6721 tcaccatatt ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc
6781 ctcccaaatt gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt
6841 tgtaggtaac caggcgcgtg aattcactcc tcaggtgcag gctgcctatc agaaggtggt
6901 ggctggtgtg gccaatgccc tggctcacaa ataccactga gatctttttc cctctgccaa
6961 aaattatggg gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt
7021 tattttcatt gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg
7081 gagggcaaat catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc
7141 ccatatgctg gctgccatga acaaaggttg gctataaaga ggtcatcagt atatgaaaca
7201 gccccctgct gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt
7261 tatattttgt tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt
7321 actagccaga tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctctt
7381 atggagatcc ctcgacctgc agcccaagct tggcgtaatc atggtcatag ctgtttcctg
7441 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
7501 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
7561 ctttccagtc gggaaacctg tcgtgccagc ggatcgacag tactaagctt tactagggac
7621 aggattggtg acagaaaagc cccatcctta ggcctcctcc ttcctagtct cctgatattg
7681 ggtctaaccc ccacctcctg ttaggcagat tccttatctg gtgacacacc cccatttcct
7741 ggagccatct ctctccttgc cagaacctct aaggtttgct tacgatggag ccagagagga
7801 tcctgggagg gagagcttgg cagggggtgg gagggaaggg ggggatgcgt gacctgcccg
7861 gttctcagtg gccaccctgc gctaccctct cccagaacct gagctgctct gacgcggctg
7921 tctggtgcgt ttcactgatc ctggtgctgc agcttcctta cacttcccaa gaggagaagc
7981 agtttggaaa aacaaaatca gaataagttg gtcctgagtt ctaactttgg ctcttcacct
8041 ttctagtccc caatttatat tgttcctccg tgcgtcagtt ttacctgtga gataaggcca
8101 gtagccagcc ccgtcctggc agggctgtgg tgaggagggg ggtgtccgtg tggaaaactc
8161 cctttgtgag aatggtgcgt cctaggtgtt caccaggtcg tggccgcctc tactcccttt
8221 ctctttctcc atccttcttt ccttaaagag tccccagtgc tatctgggac atattcctcc
8281 gcccagagca gggtcccgct tccctaaggc cctgctctgg gcttctgggt ttgagtcctt
8341 ggcaagccca ggagaggcgc tcaggcttcc ctgtccccct tcctcgtcca ccatctcatg
8401 cccctggctc tcctgcccct tccctacagg ggttcctggc tctgctctaa gggcaagggc
8461 gaattcgcgg ccgctaaatt caattcgccc tatagtgagt cgtattacaa ttcactggcc
8521 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca
8581 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc
8641 caacagttgc gcagcctata cgtacggcag tttaaggttt acacctataa aagagagagc
8701 cgttatcgtc tgtttgtgga tgtacagagt gatattattg acacgccggg gcgacggatg
8761 gtgatccccc tggccagtgc acgtctgctg tcagataaag tctcccgtga actttacccg
8821 gtggtgcata tcggggatga aagctggcgc atgatgacca ccgatatggc cagtgtgccg
8881 gtctccgtta tcggggaaga agtggctgat ctcagccacc gcgaaaatga catcaaaaac
8941 gccattaacc tgatgttctg gggaatataa atgtcaggca tgagattatc aaaaaggatc
9001 ttcacctaga tccttttcac gtagaaagcc agtccgcaga aacggtgctg accccggatg
9061 aatgtcagct actgggctat ctggacaagg gaaaacgcaa gcgcaaagag aaagcaggta
9121 gcttgcagtg ggcttacatg gcgatagcta gactgggcgg ttttatggac agcaagcgaa
9181 ccggaattgc cagctggggc gccctctggt aaggttggga agccctgcaa agtaaactgg
9241 atggctttct tgccgccaag gatctgatgg cgcaggggat caagctctga tcaagagaca
9301 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
9361 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
9421 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
9481 ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc
9541 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
9601 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
9661 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
9721 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
9781 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
9841 aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atgcatggcg atgcctgctt
9901 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
9961 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
10021 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
10081 catcgccttc tatcgccttc ttgacgagtt cttctgaatt attaacgctt acaatttcct
10141 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatca ggtggcactt
10201 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
10261 atccgctcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
10321 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
10381 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
10441 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
10501 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
10561 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
10621 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
10681 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
10741 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
10801 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
10861 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
10921 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
10981 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
11041 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
11101 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
11161 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
11221 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
11281 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
11341 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
11401 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
11461 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
11521 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
11581 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
11641 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
11701 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
11761 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
11821 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
11881 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
11941 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
12001 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
12061 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
12121 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
12181 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
12241 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
12301 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctca
12361 gaattaaccc tcactaaagg gt
//
