| 质粒名称: | pEF1a-BirA-V5-neo |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 100548 |
| 存储实验室: | Jian Xu |
|
载体骨架基本信息 |
|
| 载体名称: | pEF1a-V5-neo |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 6174 bp |
| 完整质粒大小: | 7088 bp |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 新霉素（G418） |
| 高拷贝/低拷贝: | -- |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | C-terminal V5-tagged BirA |
| 物种来源: | E. coli |
| 插入大小: | 963 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | Human EF1a |
| 标签/融合蛋白: | V5，His（插入C端子） |
| 克隆方法: | Gibson克隆 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | TAATACGACTCACTATAGGG |
| 3' 测序引物 | TAGAAGGCACAGTCGAGG |
|
文献引用方法 |
|
| 材料和方法部分: | pEF1a-BirA-V5-neo was a gift from Jian Xu (Addgene plasmid # 100548 ; http://n2t.net/addgene:100548 ; RRID:Addgene_100548) |
| 参考文献部分: | In Situ Capture of Chromatin Interactions by Biotinylated dCas9. Liu X, Zhang Y, Chen Y, Li M, Zhou F, Li K, Cao H, Ni M, Liu Y, Gu Z, Dickerson KE, Xie S, Hon GC, Xuan Z, Zhang MQ, Shao Z, Xu J. Cell. 2017 Aug 24;170(5):1028-1043.e19. doi: 10.1016/j.cell.2017.08.003. 10.1016/j.cell.2017.08.003 PubMed 28841410 |
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 pEF1a-BirA-V5-neo 7088 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Expresses C-terminal V5-tagged BirA biotin ligase.
ACCESSION .
VERSION .
KEYWORDS pEF1a-BirA-V5-neo
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7088)
AUTHORS Liu X, Zhang Y, Chen Y, Li M, Zhou F, Li K, Cao H, Ni M, Liu Y, Gu
Z, Dickerson KE, Xie S, Hon GC, Xuan Z, Zhang MQ, Shao Z, Xu J
TITLE In Situ Capture of Chromatin Interactions by Biotinylated dCas9.
JOURNAL Cell. 2017 Aug 24;170(5):1028-1043.e19. doi:
10.1016/j.cell.2017.08.003.
PUBMED 28841410
REFERENCE 2 (bases 1 to 7088)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7088
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS 18..59
/codon_start=1
/product="epitope tag from simian virus 5"
/label=V5 tag
/translation="GKPIPNPLLGLDST"
CDS 69..86
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(109..126)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 115..339
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 385..813
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(472..491)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 682..703
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(822..842)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 827..1156
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 1007..1142
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 1069..1088
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 1223..2017
/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(1277..1296)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 1887..1906
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 2193..2314
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2230..2249)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 2284..2303
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(2363..2379)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(2363..2379)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(2376..2398)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 2387..2403
/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(2411..2441)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2456..2477
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(2594..2611)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2754..3342)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2834..2853)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(3513..4373)
/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 4136..4155
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(4374..4478)
/gene="bla"
/label=AmpR promoter
primer_bind complement(4553..4572)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
promoter 4860..6038
/label=EF-1-alpha promoter
/note="strong constitutive promoter for human elongation
factor EF-1-alpha"
intron 5091..6029
/label=EF-1-alpha intron A
/note="intron upstream of the start codon of human
EF-1-alpha"
primer_bind 5986..6006
/label=EF1a-F
/note="Human elongation factor-1a promoter, forward primer"
primer_bind 6055..6074
/label=T7
/note="T7 promoter, forward primer"
promoter 6055..6073
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 6125..7087
/codon_start=1
/gene="birA"
/product="E. coli biotin protein ligase"
/label=BirA
/translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
ORIGIN
1 ctagagggcc cttcgaaggt aagcctatcc ctaaccctct cctcggtctc gattctacgc
61 gtaccggtca tcatcaccat caccattgag tttaaacccg ctgatcagcc tcgactgtgc
121 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
181 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta
241 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
301 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
361 gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg
421 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
481 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
541 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
601 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
661 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
721 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
781 atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg
841 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
901 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
961 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
1021 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
1081 ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
1141 cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga
1201 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
1261 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
1321 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
1381 gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac
1441 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
1501 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
1561 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
1621 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
1681 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
1741 gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
1801 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
1861 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
1921 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
1981 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgcgaaa
2041 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
2101 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
2161 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
2221 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
2281 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
2341 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
2401 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
2461 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
2521 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
2581 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
2641 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
2701 agaacatgtg agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
2761 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
2821 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
2881 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
2941 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
3001 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
3061 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
3121 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
3181 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
3241 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
3301 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
3361 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
3421 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
3481 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
3541 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
3601 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
3661 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
3721 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
3781 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
3841 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
3901 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
3961 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
4021 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
4081 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
4141 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
4201 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
4261 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
4321 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
4381 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
4441 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
4501 cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
4561 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
4621 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
4681 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
4741 gttgacattg attattgact aggcttttgc aaaaagcttt gcaaagatgg ataaagtttt
4801 aaacagagag gaatctttgc agctaatgga ccttctaggt cttgaaagga gtgggaattg
4861 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
4921 gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
4981 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
5041 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg
5101 tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta
5161 cttccacctg gctgcagtac gtgattcttg atcccgagct tcgggttgga agtgggtggg
5221 agagttcgag gccttgcgct taaggagccc cttcgcctcg tgcttgagtt gaggcctggc
5281 ctgggcgctg gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt
5341 tcgataagtc tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc
5401 aagatagtct tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt tttggggccg
5461 cgggcggcga cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag
5521 cgcggccacc gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg
5581 gcctcgcgcc gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag
5641 ttgcgtgagc ggaaagatgg ccgcttcccg gccctgctgc agggagctca aaatggagga
5701 cgcggcgctc gggagagcgg gcgggtgagt cacccacaca aaggaaaagg gcctttccgt
5761 cctcagccgt cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt
5821 agttctcgag cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg
5881 agtttcccca cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat
5941 tctccttgga atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag
6001 tggttcaaag tttttttctt ccatttcagg tgtcgtgagg aattagcttg gtactaatac
6061 gactcactat agggagaccc aagctggcta ggtaagcttg gtaccgagct cggatccgcc
6121 agccatgaag gataacaccg tgccactgaa attgattgcc ctgttagcga acggtgaatt
6181 tcactctggc gagcagttgg gtgaaacgct gggaatgagc cgggcggcta ttaataaaca
6241 cattcagaca ctgcgtgact ggggcgttga tgtctttacc gttccgggta aaggatacag
6301 cctgcctgag cctatccagt tacttaatgc taaacagata ttgggtcagc tggatggcgg
6361 tagtgtagcc gtgctgccag tgattgactc cacgaatcag taccttcttg atcgtatcgg
6421 agagcttaaa tcgggcgatg cttgcattgc agaataccag caggctggcc gtggtcgccg
6481 gggtcggaaa tggttttcgc cttttggcgc aaacttatat ttgtcgatgt tctggcgtct
6541 ggaacaaggc ccggcggcgg cgattggttt aagtctggtt atcggtatcg tgatggcgga
6601 agtattacgc aagctgggtg cagataaagt tcgtgttaaa tggcctaatg acctctatct
6661 gcaggatcgc aagctggcag gcattctggt ggagctgact ggcaaaactg gcgatgcggc
6721 gcaaatagtc attggagccg ggatcaacat ggcaatgcgc cgtgttgaag agagtgtcgt
6781 taatcagggg tggatcacgc tgcaggaagc ggggatcaat ctcgatcgta atacgttggc
6841 ggccatgcta atacgtgaat tacgtgctgc gttggaactc ttcgaacaag aaggattggc
6901 accttatctg tcgcgctggg aaaagctgga taattttatt aatcgcccag tgaaacttat
6961 cattggtgat aaagaaatat ttggcatttc acgcggaata gacaaacagg gggctttatt
7021 acttgagcag gatggaataa taaaaccctg gatgggcggt gaaatatccc tgcgtagtgc
7081 agaaaaat
//
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