| 质粒名称: | pCMV-T7-hAcrVA1-NLS(sv40) (BPK5050) |
|---|---|
| 出品公司: | addgene |
| 目录编号: | 115136 |
| 存储实验室: | Joseph Bondy-Denomy, Keith Joung, Benjamin Kleinstiver |
|
载体骨架基本信息 |
|
| 载体名称: | JDS246 |
| 载体出品公司: | Joung lab (Addgene Plasmid #43861) |
| 完整质粒大小: | 3960 bp |
| 载体类型: | 哺乳细胞表达 |
| 载体抗性: | 氨苄霉素 |
| 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: | human codon optimized AcrVA1 anti-CRISPR protein / BPK5050 |
| 物种来源: | Synthetic |
| 标签/融合蛋白: | NLS(SV40) (C terminal on insert) |
| 克隆方法: | Gibson Cloning |
| 5’测序引物 | CMV-F |
| 3’测序引物 | BGH-rev |
|
文献引用方法 |
|
| 材料和方法部分: | pCMV-T7-hAcrVA1-NLS(sv40) (BPK5050) was a gift from Joseph Bondy-Denomy & Keith Joung & Benjamin Kleinstiver (Addgene plasmid # 115136 ; http://n2t.net/addgene:115136 ; RRID:Addgene_115136) |
| 参考文献部分: | Discovery of widespread Type I and Type V CRISPR-Cas inhibitors. Marino ND, Zhang JY, Borges AL, Sousa AA, Leon LM, Rauch BJ, Walton RT, Berry JD, Joung JK, Kleinstiver BP, Bondy-Denomy J. Science. 2018 Sep 6. pii: science.aau5174. doi: 10.1126/science.aau5174. 10.1126/science.aau5174 PubMed 30190308 |
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
 (BPK5050)质粒图谱.png)
载体简介
载体序列
LOCUS pCMV-T7-hAcrVA1-NLS(sv40) (BPK5050) 3960 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION CMV-T7 promoter expression plasmid for human codon optimized AcrVA1
anti-CRISPR protein with C-terminal NLS .
ACCESSION .
VERSION .
KEYWORDS pCMV-T7-hAcrVA1-NLS(sv40) (BPK5050)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 3960)
AUTHORS Marino ND, Zhang JY, Borges AL, Sousa AA, Leon LM, Rauch BJ, Walton
RT, Berry JD, Joung JK, Kleinstiver BP, Bondy-Denomy J
TITLE Discovery of widespread Type I and Type V CRISPR-Cas inhibitors.
JOURNAL Science. 2018 Sep 6. pii: science.aau5174. doi:
10.1126/science.aau5174.
PUBMED 30190308
REFERENCE 2 (bases 1 to 3960)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..3960
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS 538..558
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 567..584
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(607..624)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 613..837
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind complement(908..924)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(908..924)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(921..943)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 932..948
/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(956..986)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 1001..1022
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(1139..1156)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(1310..1898)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(1390..1409)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(2069..2929)
/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 2692..2711
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(2930..3034)
/gene="bla"
/label=AmpR promoter
primer_bind complement(3113..3132)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 3304..3683
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 3684..3887
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 3838..3858
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 3929..3948
/label=T7
/note="T7 promoter, forward primer"
promoter 3929..3947
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
regulatory 3955..4
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
ORIGIN
1 atgggctatg aggcaaaaga acggtacgct aaaaagaaga tgcaggagaa tacaaaaata
61 gacacactta cagacgagca gcacgacgca ctggcccaac tgtgtgcctt ccgacataaa
121 tttcactcca ataaggacag tctcttcctc tctgagtccg cttttagcgg tgagttttcc
181 tttgagatgc aaagcgatga aaatagcaag ctgagagaag taggtcttcc cacaatagaa
241 tggtcattct acgacaacag tcacattccg gacgatagct tccgagagtg gtttaacttt
301 gcaaactact cagagctctc tgagacaata caagaacagg gtctggagct ggacctcgac
361 gacgatgaga catacgagct ggtttatgat gaactctata cagaggctat gggagaatat
421 gaagagctta atcaagatat tgagaagtat ctgaggagga ttgacgagga acatgggacc
481 cagtattgcc caaccgggtt tgcccgcctt cgaggatccg gcggtggagg cagtgggccc
541 aagaagaaga ggaaagtctg accggtcatc atcaccatca ccattgagtt taaacccgct
601 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc
661 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg
721 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca
781 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt
841 ctgaggcgga aagaaccagc tggggctcga taccgtcgac ctctagctag agcttggcgt
901 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
961 tacgagccgg aagcataaag tgtaaagcct agggtgccta atgagtgagc taactcacat
1021 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
1081 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
1141 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
1201 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
1261 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
1321 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
1381 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
1441 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
1501 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
1561 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
1621 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
1681 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
1741 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
1801 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
1861 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
1921 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
1981 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
2041 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
2101 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
2161 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
2221 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
2281 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
2341 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
2401 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
2461 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
2521 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
2581 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
2641 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
2701 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
2761 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
2821 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
2881 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
2941 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
3001 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
3061 acgtcgacgg atcgggagat cgatctcccg atcccctagg gtcgactctc agtacaatct
3121 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
3181 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
3241 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
3301 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
3361 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
3421 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
3481 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
3541 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
3601 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
3661 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
3721 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
3781 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
3841 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctggt ttagtgaacc
3901 gtcagatccg ctagagatcc gcggccgcta atacgactca ctatagggag agccgccacc
//
