| 质粒名称: | pJH376 |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 86842 |
| 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: | 5684 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: | eacrIIA4 / LMOG_02993 |
| 物种来源: | Listeria monocytogenes |
| 启动子: | CMV |
| 克隆方法: | Gibson Cloning |
| 5’测序引物 | CMV-F |
| 3’测序引物 | SV40pA-R |
|
文献引用方法 |
|
| 材料和方法部分: | pJH376 was a gift from Joseph Bondy-Denomy (Addgene plasmid # 86842 ; http://n2t.net/addgene:86842 ; RRID:Addgene_86842) |
| 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 pJH376 5684 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION mamalian expression of AcrIIA4.
ACCESSION .
VERSION .
KEYWORDS pJH376
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5684)
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 5684)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..5684
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 25..404
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 405..608
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 559..579
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 653..672
/label=T7
/note="T7 promoter, forward primer"
promoter 653..671
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(1065..1082)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 1071..1295
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 1341..1769
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1428..1447)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 1638..1659
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(1778..1798)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 1783..2112
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 1963..2098
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 2025..2044
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 2179..2973
/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(2233..2252)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 2843..2862
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 3147..3268
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(3184..3203)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 3238..3257
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(3317..3333)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(3317..3333)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(3330..3352)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 3341..3357
/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(3365..3395)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 3410..3431
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(3548..3565)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3719..4307)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3799..3818)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(4478..5338)
/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 5101..5120
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(5339..5443)
/gene="bla"
/label=AmpR promoter
primer_bind complement(5518..5537)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa
61 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
121 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
181 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
241 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
301 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
361 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
421 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
481 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
541 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
601 gcagagctct ctggctaact agagaaccca ctgcttactg gcttatcgaa attaatacga
661 ctcactatag ggagacccaa gctggctagc gtttaaactt aagcttggta ccgagctcgg
721 atccgccacc atgaacatta acgacctcat acgagagatt aagaacaaag attacaccgt
781 caaactgtca ggaactgata gtaactcaat cacccagctt attatcaggg taaacaatga
841 tgggaatgaa tatgtgatat ctgagagcga aaacgagtct atcgtcgaga aattcatttc
901 cgcttttaag aacgggtgga atcaggaata tgaggatgaa gaagaatttt acaatgacat
961 gcagacgatc acgttgaaaa gtgaactgaa ctaagaattc tgcagatatc cagcacagtg
1021 gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga ctgtgccttc
1081 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
1141 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
1201 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
1261 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg
1321 gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
1381 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
1441 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
1501 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
1561 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
1621 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
1681 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
1741 gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt
1801 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
1861 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
1921 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
1981 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
2041 gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
2101 ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca
2161 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
2221 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
2281 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
2341 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
2401 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
2461 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
2521 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
2581 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
2641 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
2701 aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
2761 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
2821 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
2881 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
2941 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
3001 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
3061 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
3121 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat
3181 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
3241 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga
3301 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
3361 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
3421 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
3481 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
3541 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
3601 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
3661 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
3721 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
3781 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
3841 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
3901 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
3961 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
4021 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
4081 acaggattag cagagcgagg tatgtaggca gtgctacaga gttcttgaag tggtggccta
4141 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
4201 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
4261 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
4321 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
4381 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
4441 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
4501 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
4561 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
4621 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
4681 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
4741 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
4801 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
4861 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
4921 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
4981 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
5041 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
5101 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
5161 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
5221 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
5281 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
5341 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
5401 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
5461 tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac
5521 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
5581 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
5641 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcg
//
