| 质粒名称: | pCMV-T7-hAcrVA2.1-NLS(sv40) (BPK5059) |
|---|---|
| 出品公司: | addgene |
| 目录编号: | 115137 |
| 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: | 4407 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 AcrVA2.1 anti-CRISPR protein / BPK5059 |
| 物种来源: | Synthetic |
| 标签/融合蛋白: | Gibson Cloning |
| 克隆方法: | NLS(SV40) (C terminal on insert) |
| 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-hAcrVA2.1-NLS(sv40) (BPK5059) was a gift from Joseph Bondy-Denomy & Keith Joung & Benjamin Kleinstiver (Addgene plasmid # 115137 ; http://n2t.net/addgene:115137 ; RRID:Addgene_115137) |
| 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 |
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pCMV-T7-hAcrVA2.1-NLS(sv40) (BPK5059)载体质粒基本信息
pCMV-T7-hAcrVA2.1-NLS(sv40) (BPK5059)质粒图谱载体图谱和pCMV-T7-hAcrVA2.1-NLS(sv40) (BPK5059)载体序列质粒序列多克隆位点信息
 (BPK5059)质粒图谱.png)
pCMV-T7-hAcrVA2.1-NLS(sv40) (BPK5059)质粒载体简介
pCMV-T7-hAcrVA2.1-NLS(sv40) (BPK5059)质粒序列载体序列
LOCUS pCMV-T7-hAcrVA2.1-NLS(sv40) (BPK5059) 4407 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION CMV-T7 promoter expression plasmid for human codon optimized
AcrVA2.1 anti-CRISPR protein with C-terminal NLS .
ACCESSION .
VERSION .
KEYWORDS pCMV-T7-hAcrVA2.1-NLS(sv40) (BPK5059)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4407)
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 4407)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..4407
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS 985..1005
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 1014..1031
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(1054..1071)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 1060..1284
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind complement(1355..1371)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1355..1371)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(1368..1390)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 1379..1395
/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(1403..1433)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 1448..1469
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(1586..1603)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(1757..2345)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(1837..1856)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(2516..3376)
/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 3139..3158
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(3377..3481)
/gene="bla"
/label=AmpR promoter
primer_bind complement(3560..3579)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 3751..4130
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 4131..4334
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 4285..4305
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 4376..4395
/label=T7
/note="T7 promoter, forward primer"
promoter 4376..4394
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
regulatory 4402..4
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
ORIGIN
1 atgggccacc acacgatagc tagaatgaac gcctttaaca aggctttcgg caatgccaag
61 gactgttaca agaaaatgca ggcctggcac ctcaataaca aaccaaaaca tatctttagc
121 ccgctccaga ataccctctc actgaatgaa ggcttggcag cgctttatga gcttcatggc
181 gggaaggagg atgaacacat tctgtctatc ctctgttgcc tgtatctgta tgggacatgg
241 agaaacacac tgggcatata ccaactggat gaagaaataa ttaaggattg caaagagttg
301 cccgatgaca cacctacaag cattttcctt aacttgcctg actggtgtgt ttacgtcgat
361 atctcaagcg cgaaaattgc caccatagat ggcggcgtcg caaagcacat aaagggtttt
421 tgggcgatct acgacaacat tgagatgcac ggcgtgaacc acgatgtcct caattttata
481 atcgacactg atacggacaa caatatctat gtaccacagt ctcttatcct cagttctgag
541 atgtcagtag cggagagtct tgactacggt ctgaccctct tcgggtatga cgaatctaac
601 gagttggtca aagggatgct tccatacctc ctttggctgt gcgttgctga gccggacata
661 acacacaaag gcttgccagt cagcagagag gagcttacaa agcctaaaca cggaataaat
721 aagaagacgg gggccttcgt gacgcctagc gaacctttca tatatcagat aggagaacgg
781 ctcggtgggg aggtgagaag ataccagagc ctgatagacg atgaaaagaa ccaaaataga
841 catcacacta aaaggccgca cataaggagg gggcactggc acggctactg gcaggggacg
901 ggacaggcaa aggagtttaa ggttcgatgg caaccggctg tttttgtaaa ctcaggtgtc
961 ggatccggcg gtggaggcag tgggcccaag aagaagagga aagtctgacc ggtcatcatc
1021 accatcacca ttgagtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
1081 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
1141 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
1201 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
1261 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctcgatac
1321 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
1381 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctagg
1441 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
1501 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
1561 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
1621 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
1681 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
1741 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
1801 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
1861 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
1921 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
1981 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
2041 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
2101 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
2161 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
2221 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
2281 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
2341 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
2401 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
2461 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
2521 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
2581 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
2641 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
2701 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
2761 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
2821 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
2881 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
2941 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
3001 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
3061 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
3121 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
3181 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
3241 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
3301 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
3361 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
3421 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
3481 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatcga tctcccgatc
3541 ccctagggtc gactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc
3601 tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa
3661 ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc
3721 ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt
3781 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
3841 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
3901 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
3961 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
4021 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
4081 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
4141 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
4201 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
4261 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
4321 atataagcag agctggttta gtgaaccgtc agatccgcta gagatccgcg gccgctaata
4381 cgactcacta tagggagagc cgccacc
//
