| 质粒名称: | pJH375 |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 86841 |
| 存储实验室: | Joseph Bondy-Denomy |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA3.1(+) |
| 完整质粒大小: | 5795 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: | acrIIA3 / Axk13_03345 |
| 物种来源: | Streptococcus pyogenes |
| 启动子: | CMV |
| 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: | pJH375 was a gift from Joseph Bondy-Denomy (Addgene plasmid # 86841 ; http://n2t.net/addgene:86841 ; RRID:Addgene_86841) |
| 参考文献部分: | Inhibition of CRISPR-Cas9 with Bacteriophage Proteins. Rauch BJ, Silvis MR, Hultquist JF, Waters CS, McGregor MJ, Krogan NJ, Bondy-Denomy J. Cell. 2016 Dec 22. pii: S0092-8674(16)31683-X. doi: 10.1016/j.cell.2016.12.009. 10.1016/j.cell.2016.12.009 PubMed 28041849 |
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 pJH375 5795 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION mamalian expression of AcrIIA3.
ACCESSION .
VERSION .
KEYWORDS pJH375
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5795)
AUTHORS Rauch BJ, Silvis MR, Hultquist JF, Waters CS, McGregor MJ, Krogan
NJ, Bondy-Denomy J
TITLE Inhibition of CRISPR-Cas9 with Bacteriophage Proteins.
JOURNAL Cell. 2016 Dec 22. pii: S0092-8674(16)31683-X. doi:
10.1016/j.cell.2016.12.009.
PUBMED 28041849
REFERENCE 2 (bases 1 to 5795)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..5795
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 27..406
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 407..610
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 561..581
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 655..674
/label=T7
/note="T7 promoter, forward primer"
promoter 655..673
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(1178..1195)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 1184..1408
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 1454..1882
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1541..1560)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 1751..1772
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(1891..1911)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 1896..2225
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 2076..2211
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 2138..2157
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 2292..3086
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(2346..2365)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 2956..2975
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 3260..3381
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(3297..3316)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 3351..3370
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(3430..3446)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(3430..3446)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(3443..3465)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 3454..3470
/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(3478..3508)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 3523..3544
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(3661..3678)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3832..4420)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3912..3931)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(4591..5451)
/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 5214..5233
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(5452..5556)
/gene="bla"
/label=AmpR promoter
primer_bind complement(5631..5650)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc
61 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
121 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
181 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
241 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
301 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
361 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
421 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
481 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
541 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
601 aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg aaattaatac
661 gactcactat agggagaccc aagctggcta gcgtttaaac ttaagcttgg taccgagctc
721 ggatccgcca ccatgacaaa gtataataag tcagagataa tgaaaaacgc atgggcgatg
781 tttaacagct atgaatggga tgttgaaaat tttaaattcg tgagtgctga gaacaagacc
841 ttctctaatt gcctcaagga agcctgggca gaagagaagg aatatgtcga gaggaaagcc
901 aaagaaacag ccgaggcgcc ccggtctgag gaggctaaag catgggactg ggcgtgcaga
961 aaattgaatg taaacgattt gcagaacatt gacgccactg acaaagtctt ctatgtggtt
1021 gacatgcaaa aagaaatgtg gacttccaat gtgtgggcgc aggcaatcaa ggctgtagaa
1081 ctgtatgtga agctcggtct tgcttaagaa ttctgcagat atccagcaca gtggcggccg
1141 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
1201 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
1261 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
1321 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
1381 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
1441 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
1501 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
1561 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
1621 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
1681 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
1741 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
1801 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
1861 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
1921 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
1981 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
2041 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
2101 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
2161 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
2221 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
2281 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
2341 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
2401 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
2461 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
2521 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
2581 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
2641 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
2701 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
2761 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
2821 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
2881 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
2941 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
3001 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
3061 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
3121 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
3181 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
3241 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
3301 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
3361 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
3421 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
3481 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
3541 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
3601 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
3661 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
3721 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
3781 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
3841 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
3901 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
3961 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
4021 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
4081 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
4141 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
4201 tagcagagcg aggtatgtag gcagtgctac agagttcttg aagtggtggc ctaactacgg
4261 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
4321 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
4381 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
4441 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
4501 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
4561 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
4621 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
4681 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
4741 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
4801 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
4861 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
4921 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
4981 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
5041 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
5101 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
5161 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
5221 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
5281 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
5341 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
5401 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
5461 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
5521 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
5581 tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag tacaatctgc
5641 tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag
5701 tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag
5761 aatctgctta gggttaggcg ttttgcgctg cttcg
//
