| 质粒名称: | pCMV-T7-hAcrVA3-NLS(sv40) (BPK5077) |
|---|---|
| 出品公司: | addgene |
| 目录编号: | 115140 |
| 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 (Plasmid #43861) |
| 载体出品公司: | Joung lab (Addgene Plasmid #43861) |
| 完整质粒大小: | 3954 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 AcrVA3 anti-CRISPR protein / BPK5077 |
| 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-hAcrVA3-NLS(sv40) (BPK5077) was a gift from Joseph Bondy-Denomy & Keith Joung & Benjamin Kleinstiver (Addgene plasmid # 115140 ; http://n2t.net/addgene:115140 ; RRID:Addgene_115140) |
| 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
 (BPK5077)质粒图谱.png)
载体简介
载体序列
LOCUS pCMV-T7-hAcrVA3-NLS(sv40) (BPK5077) 3954 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION CMV-T7 promoter expression plasmid for human codon optimized AcrVA3
anti-CRISPR protein with C-terminal NLS .
ACCESSION .
VERSION .
KEYWORDS pCMV-T7-hAcrVA3-NLS(sv40) (BPK5077)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 3954)
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 3954)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..3954
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS 532..552
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 561..578
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(601..618)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 607..831
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind complement(902..918)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(902..918)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(915..937)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 926..942
/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(950..980)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 995..1016
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(1133..1150)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(1304..1892)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(1384..1403)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(2063..2923)
/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 2686..2705
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(2924..3028)
/gene="bla"
/label=AmpR promoter
primer_bind complement(3107..3126)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 3298..3677
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 3678..3881
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 3832..3852
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 3923..3942
/label=T7
/note="T7 promoter, forward primer"
promoter 3923..3941
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
regulatory 3949..4
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
ORIGIN
1 atgggcgttg gtaaatcaaa aattgactgg caatctatag actggactaa aaccaatgcg
61 caaattgccc aagaatgtgg gcgagcatat aacaccgtgt gcaaaatgcg gggtaaactt
121 ggcaagagcc atcaaggcgc caaatctccg cgcaaggata aaggcatcag tcgccctcaa
181 ccccatttga accggcttga gtaccaggca cttgccacag cgaaagcaaa ggccagccca
241 aaagcgggac gattcgagac aaacacaaaa gcaaaaacat ggactctgaa atcacctgat
301 aataaaacat atacgtttac gaatctgatg cactttgttc gcacaaaccc acatctgttt
361 gaccccgatg atgtagtgtg gcggacaaaa agcaatggtg tggaatggtg cagggcctca
421 agcgggctgg cgctgcttgc caaacgcaag aaagcgccac tttcttggaa aggttggcgc
481 cttatttctc tcacaaaaga taacaaagga tccggcggtg gaggcagtgg gcccaagaag
541 aagaggaaag tctgaccggt catcatcacc atcaccattg agtttaaacc cgctgatcag
601 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
661 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
721 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
781 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
841 cggaaagaac cagctggggc tcgataccgt cgacctctag ctagagcttg gcgtaatcat
901 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
961 ccggaagcat aaagtgtaaa gcctagggtg cctaatgagt gagctaactc acattaattg
1021 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
1081 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
1141 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
1201 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
1261 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
1321 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
1381 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
1441 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
1501 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
1561 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
1621 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
1681 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
1741 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
1801 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
1861 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
1921 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
1981 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
2041 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
2101 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
2161 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
2221 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
2281 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
2341 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
2401 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
2461 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
2521 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
2581 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
2641 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
2701 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
2761 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
2821 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
2881 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
2941 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
3001 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
3061 acggatcggg agatcgatct cccgatcccc tagggtcgac tctcagtaca atctgctctg
3121 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
3181 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
3241 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
3301 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
3361 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
3421 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
3481 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
3541 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
3601 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
3661 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
3721 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
3781 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
3841 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg aaccgtcaga
3901 tccgctagag atccgcggcc gctaatacga ctcactatag ggagagccgc cacc
//
