| 质粒名称: | pAAV_LP1B_St1Cas9_LMD-9_SpA_U6_sgRNA |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 110624 |
| 存储实验室: | Yannick Doyon |
|
载体骨架基本信息 |
|
| 载体名称: | pX602 |
| 空载体大小: | 6743 bp |
| 完整质粒大小: | 6743 bp |
| 载体类型: | 哺乳、 Mouse Targeting, AAV, CRISPR细胞表达 |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | NEB Stable |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称1: | St1Cas9 LMD-9 |
| 物种来源: | H. sapiens (human), Synthetic |
| 启动子: | LP1B |
| 标签/融合蛋白: |
SV40 NLS (N terminal on insert)
SV40 NLS (C terminal on insert)
|
| 克隆方法: | Gibson Cloning |
| 5’测序引物： | U6 |
| 插入基因名称2: | sgRNA for St1Cas9 |
| 物种来源: | H. sapiens (human), Synthetic |
| 启动子: | hU6 |
| 克隆方法: | Gibson Cloning |
| 5’测序引物： | U6 |
|
文献引用方法 |
|
| 材料和方法部分: | pAAV_LP1B_St1Cas9_LMD-9_SpA_U6_sgRNA was a gift from Yannick Doyon (Addgene plasmid # 110624 ; http://n2t.net/addgene:110624 ; RRID:Addgene_110624) |
| 参考文献部分: | Versatile and robust genome editing with Streptococcus thermophilus CRISPR1-Cas9. Agudelo D, Carter S, Velimirovic M, Duringer A, Rivest JF, Levesque S, Loehr J, Mouchiroud M, Cyr D, Waters PJ, Laplante M, Moineau S, Goulet A, Doyon Y. Genome Res. 2020 Jan;30(1):107-117. doi: 10.1101/gr.255414.119. Epub 2020 Jan 3. 10.1101/gr.255414.119 PubMed 31900288 |
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 pAAV_LP1B_St1Cas9_LMD-9_SpA_U6_sgRNA 6743 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION A single vector AAV-Cas9 system containing a liver-specific promoter
with Cas9 from Streptococcus thermophilus CRISPR1 (St1Cas9 LMD-9)
and its U6-driven sgRNA.
ACCESSION .
VERSION .
KEYWORDS pAAV_LP1B_St1Cas9_LMD-9_SpA_U6_sgRNA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6743)
AUTHORS Agudelo D, Carter S, Velimirovic M, Duringer A, Rivest JF, Levesque
S, Loehr J, Mouchiroud M, Cyr D, Waters PJ, Laplante M, Moineau S,
Goulet A, Doyon Y
TITLE Versatile and robust genome editing with Streptococcus thermophilus
CRISPR1-Cas9.
JOURNAL Genome Res. 2020 Jan;30(1):107-117. doi: 10.1101/gr.255414.119. Epub
2020 Jan 3.
PUBMED 31900288
REFERENCE 2 (bases 1 to 6743)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..6743
/organism="synthetic DNA construct"
/mol_type="other DNA"
repeat_region 61..190
/label=AAV2 ITR (alternate)
/note="Functional equivalent of wild-type AAV2 ITR"
regulatory 723..732
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 738..758
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 783..4142
/codon_start=1
/product="Cas9 endonuclease from the Streptococcus
thermophilus Type II CRISPR/Cas system (Esvelt et al.,
2013)"
/label=St1Cas9
/note="generates RNA-guided double strand breaks in DNA"
/translation="SDLVLGLDIGIGSVGVGILNKVTGEIIHKNSRIFPAAQAENNLVR
RTNRQGRRLARRKKHRRVRLNRLFEESGLITDFTKISINLNPYQLRVKGLTDELSNEEL
FIALKNMVKHRGISYLDDASDDGNSSVGDYAQIVKENSKQLETKTPGQIQLERYQTYGQ
LRGDFTVEKDGKKHRLINVFPTSAYRSEALRILQTQQEFNPQITDEFINRYLEILTGKR
KYYHGPGNEKSRTDYGRYRTSGETLDNIFGILIGKCTFYPDEFRAAKASYTAQEFNLLN
DLNNLTVPTETKKLSKEQKNQIINYVKNEKAMGPAKLFKYIAKLLSCDVADIKGYRIDK
SGKAEIHTFEAYRKMKTLETLDIEQMDRETLDKLAYVLTLNTEREGIQEALEHEFADGS
FSQKQVDELVQFRKANSSIFGKGWHNFSVKLMMELIPELYETSEEQMTILTRLGKQKTT
SSSNKTKYIDEKLLTEEIYNPVVAKSVRQAIKIVNAAIKEYGDFDNIVIEMARETNEDD
EKKAIQKIQKANKDEKDAAMLKAANQYNGKAELPHSVFHGHKQLATKIRLWHQQGERCL
YTGKTISIHDLINNSNQFEVDHILPLSITFDDSLANKVLVYATANQEKGQRTPYQALDS
MDDAWSFRELKAFVRESKTLSNKKKEYLLTEEDISKFDVRKKFIERNLVDTRYASRVVL
NALQEHFRAHKIDTKVSVVRGQFTSQLRRHWGIEKTRDTYHHHAVDALIIAASSQLNLW
KKQKNTLVSYSEDQLLDIETGELISDDEYKESVFKAPYQHFVDTLKSKEFEDSILFSYQ
VDSKFNRKISDATIYATRQAKVGKDKADETYVLGKIKDIYTQDGYDAFMKIYKKDKSKF
LMYRHDPQTFEKVIEPILENYPNKQINEKGKEVPCNPFLKYKEEHGYIRKYSKKGNGPE
IKSLKYYDSKLGNHIDITPKDSNNKVVLQSVSPWRADVYFNKTTGKYEILGLKYADLQF
EKGTGTYKISQEKYNDIKKKEGVDSDSEFKFTLYKNDLLLVKDTETKEQQLFRFLSRTM
PKQKHYVELKPYDKQKFEGGEALIKVLGNVANSGQCKKGLGKSNISIYKVRTDVLGNQH
IIKNEGDKPKLDF"
CDS 4155..4175
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
polyA_signal 4206..4254
/note="synthetic polyadenylation signal"
promoter complement(4378..4618)
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind complement(4428..4447)
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
primer_bind complement(4598..4618)
/label=hU6-F
/note="Human U6 promoter, forward primer"
repeat_region 4647..4787
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 4647..4776
/label=AAV2 ITR
primer_bind complement(4949..4968)
/label=F1ori-R
/note="F1 origin, reverse primer"
promoter 5018..5122
/gene="bla"
/label=AmpR promoter
CDS 5123..5983
/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(5341..5360)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 6154..6742
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6643..6662
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
61 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
121 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
181 aggggttcct gcggcctcta gactcgagcc ctaaaatggg caaacattgc aagcagcaaa
241 cagcaaacac acagccctcc ctgcctgctg accttggagc tggggcagag gtcagagacc
301 tctctgggcc catgccacct ccaacatcca ctcgacccct tggaatttcg gtggagagga
361 gcagaggttg tcctggcgtg gtttaggtag tgtgagaggg tggacacagg acgctgtggt
421 ttctgagcca gggggcgact cagatcccag ccagtggact tagcccctgt ttgctcctcc
481 gataactggg gtgaccttgg ttaatattca ccagcagcct cccccgttgc ccctctggat
541 ccactgctta aatacggacg aggacagggc cctgtctcct cagcttcagg caccaccact
601 gacctgggac agtgaatccg gactctaagg taaatataaa atttttaagt gtataatgtg
661 ttaaactact gattctaatt gtttctctct tttagattcc aacctttgga actgaaccgg
721 tcgccaccat gggcgcccca aagaagaagc ggaaggtcgg tatccacgga gtcccagcag
781 ccagcgacct ggtgctgggc ctggacatcg gcatcggcag cgtgggcgtg ggcatcctga
841 acaaggtgac cggcgagatc atccacaaga acagtcgcat cttccctgct gctcaggctg
901 agaacaacct ggtgcgccgc accaaccgcc agggtcgccg gcttgctcgc cgcaagaagc
961 accggcgcgt gcgcctgaac cgcctgttcg aggagagcgg cctgatcacc gacttcacca
1021 agatcagcat caacctgaac ccctaccagc tgcgcgtgaa gggcctgacc gacgagctga
1081 gcaacgagga gctgttcatc gccctgaaga acatggtgaa gcaccgcggc atcagctacc
1141 tggacgacgc cagcgacgac ggcaacagca gcgtgggcga ctacgcccag atcgtgaagg
1201 agaacagcaa gcagctggag accaagaccc ccggccagat ccagctggag cgctaccaga
1261 cctacggcca gctgcgcggc gacttcaccg tggagaagga cggcaagaag caccgcctga
1321 tcaacgtgtt ccccaccagc gcctaccgca gcgaggccct gcgcatcctg cagacccagc
1381 aggagttcaa cccccagatc accgacgagt tcatcaaccg ctacctggag atcctgaccg
1441 gcaagcgcaa gtactaccac ggccccggca acgagaagag ccgcaccgac tacggccgct
1501 accgcaccag cggcgagacc ctggacaaca tcttcggcat cctgatcggc aagtgcacct
1561 tctaccccga cgagttccgc gccgccaagg ccagctacac cgcccaggag ttcaatctgc
1621 tgaacgacct gaacaacctg accgtgccca ccgagaccaa gaagctgagc aaggagcaga
1681 agaaccagat catcaactac gtgaagaacg agaaggctat gggccccgcc aagctgttca
1741 agtacatcgc caagctgctg agctgcgacg tggccgacat caagggctac cgcatcgaca
1801 agagcggcaa ggccgagatc cacaccttcg aggcctaccg caagatgaag accctggaga
1861 ccctggacat cgagcagatg gaccgagaga ccctggacaa gctggcctac gtgctgaccc
1921 tgaacaccga gcgcgagggc atccaggagg ccctggagca cgagttcgcc gacggcagct
1981 tcagccagaa acaggtggac gagctggtgc agttccgcaa ggccaacagc agcatcttcg
2041 gcaagggctg gcacaacttc agcgtgaagc tgatgatgga gctgatcccc gagctgtacg
2101 agaccagcga ggagcagatg accatcctga cccgcctggg caagcagaag accaccagca
2161 gcagcaacaa gaccaagtac atcgacgaga agctgctgac cgaggagatc tacaaccccg
2221 tggtggccaa gagcgtgcgc caggccatca agatcgtgaa cgccgccatc aaggagtacg
2281 gcgacttcga caacatcgtg atcgagatgg cccgcgagac caacgaggac gacgagaaga
2341 aggccatcca gaagatccag aaggccaaca aggacgagaa ggacgccgcc atgctgaagg
2401 ccgccaacca gtacaacggc aaggccgagc tgccccacag cgtgttccac ggccacaagc
2461 agctggccac caagatccgc ctgtggcacc agcagggcga gcgctgcctg tacaccggca
2521 agaccatcag catccacgac ctgatcaaca acagcaacca gttcgaggtg gaccacatcc
2581 tgcccctgag catcaccttc gacgacagcc tggccaacaa ggtgctggtg tacgccaccg
2641 ccaaccagga gaagggccag cgcaccccct accaggccct ggacagcatg gacgacgcct
2701 ggagcttccg cgagctgaag gccttcgtgc gcgagagcaa gaccctgagc aacaagaaga
2761 aggagtatct gctgaccgag gaggacatca gcaagttcga cgtgcgcaag aagttcatcg
2821 agcgcaacct ggtggacacc cgctacgcca gccgcgtggt gctgaacgcc ctgcaggagc
2881 acttccgcgc ccacaagatc gacaccaagg tgagcgtggt gcgcggccag ttcaccagcc
2941 agctgcgccg ccactggggc atcgagaaga cccgcgacac ctaccaccac cacgccgtgg
3001 acgccctgat cattgcggct tctagccagc tgaacctgtg gaagaagcag aagaacaccc
3061 tggtgagcta cagcgaggac cagctgctgg acatcgagac cggcgagctg atcagcgacg
3121 acgagtacaa ggagagcgtg ttcaaggccc cctaccagca cttcgtggac accctgaaga
3181 gcaaggagtt cgaggacagc atcctgttca gctaccaggt ggacagcaag ttcaaccgca
3241 agatcagcga cgccaccatc tacgccaccc gccaggccaa ggtgggcaag gacaaggccg
3301 acgagaccta cgtgctgggc aagatcaagg acatctacac ccaggacggc tacgacgcct
3361 tcatgaagat ctacaagaag gacaagagca agttcctgat gtaccgccac gacccccaga
3421 ccttcgagaa ggtgatcgag cccatcctgg agaactaccc caacaagcag atcaacgaga
3481 aaggcaagga ggtgccctgc aaccccttcc tgaagtacaa ggaggagcac ggctacatcc
3541 gcaagtacag caagaagggc aacggccccg agatcaagag cctgaagtac tacgacagca
3601 agctgggcaa ccacatcgac atcaccccca aggacagcaa caacaaggtg gtgctgcaga
3661 gcgtgagccc ctggcgcgcc gacgtgtact tcaacaagac caccggcaag tacgagatcc
3721 tggggctgaa gtacgccgat ctgcagtttg agaaaggcac aggcacctac aagatcagcc
3781 aggagaagta caacgacatc aagaagaagg agggcgtgga cagcgacagc gagttcaagt
3841 tcaccctgta caagaacgac cttctgctgg tgaaggacac cgagaccaag gagcaacagc
3901 tgttccgctt cctgagccgc accatgccca agcagaagca ctacgtggag ctgaagccct
3961 acgacaagca gaagttcgag ggcggcgagg ccctgatcaa ggtgctgggc aacgtggcca
4021 acagcggcca gtgcaagaag ggcctgggca agagcaacat cagcatctac aaggtgcgca
4081 ccgacgtgct gggcaaccag cacatcatca agaacgaggg cgacaagccc aagttggact
4141 tcagcagggc tgaccccaag aagaagagga aggtgggatc ctgataagaa ttcgtcgaca
4201 agcttaataa aagatcttta ttttcattag atctgtgtgt tggttttttg tgtgcgtacg
4261 aaaaaaaaca ccctgccata aaatgacagg gtgttgattt cggcatgaag ccttatcttt
4321 gtagcttctg gtaccagagt acaaagaccg agacgcatta atcgtctcgg gtgtttcgtc
4381 ctttccacaa gatatataaa gccaagaaat cgaaatactt tcaagttacg gtaagcatat
4441 gatagtccat tttaaaacat aattttaaaa ctgcaaacta cccaagaaat tattactttc
4501 tacgtcacgt attttgtact aatatctttg tgtttacagt caaattaatt ctaattatct
4561 ctctaacagc cttgtatcgt atatgcaaat atgaaggaat catgggaaat aggccctctt
4621 cctgcccgac ctactagtgc ggccgcagga acccctagtg atggagttgg ccactccctc
4681 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt
4741 tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg
4801 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag
4861 tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
4921 gctacacttg ccagcgcctt agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
4981 acgttcgccg gcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
5041 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
5101 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
5161 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
5221 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
5281 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
5341 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
5401 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
5461 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
5521 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
5581 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
5641 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
5701 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
5761 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
5821 tggagccggt gagcgtggaa gccgcggtat cattgcagca ctggggccag atggtaagcc
5881 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
5941 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
6001 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
6061 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
6121 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
6181 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
6241 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
6301 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
6361 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
6421 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
6481 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
6541 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
6601 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
6661 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
6721 aggggggcgg agcctatgga aaa
//
