| 质粒名称: | pCMV-dCpf1-BE-YEE |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 107687 |
| 存储实验室: | Jia Chen |
|
载体骨架基本信息 |
|
| 载体名称: | CMV |
| 完整质粒大小: | 8166bp |
| 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: |
Gene/Insert name / pCMV-SV40NLS-APOBEC1(W90Y/R126E/R132E)-XTEN-dLbCpf1(D832A/E1006A/D1125A)-SV40NLS-SGGS-UGI-SV40NLS
|
| 物种来源: | R. norvegicus (rat); Lachnospiraceae bacterium Bacteriophage PBS2 |
| 插入大小: | 4764bp |
| 基因突变: | APOBEC1(W90Y/R126E/R132E), dLbCpf1(D832A/E925A/D1148A) |
| 启动子: | CMV |
| 克隆方法: | Gibson Cloning |
| 5’测序引物 | CMV-F |
| 3’测序引物 | BGH poly reverse |
|
文献引用方法 |
|
| 材料和方法部分: | pCMV-dCpf1-BE-YEE was a gift from Jia Chen (Addgene plasmid # 107687 ; http://n2t.net/addgene:107687 ; RRID:Addgene_107687) |
| 参考文献部分: | Base editing with a Cpf1-cytidine deaminase fusion. Li X, Wang Y, Liu Y, Yang B, Wang X, Wei J, Lu Z, Zhang Y, Wu J, Huang X, Yang L, Chen J. Nat Biotechnol. 2018 Mar 19. pii: nbt.4102. doi: 10.1038/nbt.4102. 10.1038/nbt.4102 PubMed 29553573 |
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 pCMV-dCpf1-BE-YEE 8166 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Expresses dLbCpf1-BE-YEE in mammalian cells.
ACCESSION .
VERSION .
KEYWORDS pCMV-dCpf1-BE-YEE
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8166)
AUTHORS Li X, Wang Y, Liu Y, Yang B, Wang X, Wei J, Lu Z, Zhang Y, Wu J,
Huang X, Yang L, Chen J
TITLE Base editing with a Cpf1-cytidine deaminase fusion.
JOURNAL Nat Biotechnol. 2018 Mar 19. pii: nbt.4102. doi: 10.1038/nbt.4102.
PUBMED 29553573
REFERENCE 2 (bases 1 to 8166)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..8166
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 6..385
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 386..589
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 540..560
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 631..650
/label=T7
/note="T7 promoter, forward primer"
promoter 631..649
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 666..686
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5112..5132
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5145..5393
/codon_start=1
/product="uracil-DNA glycosylase inhibitor from a Bacillus
subtilis bacteriophage (Mol et al., 1995)"
/label=UGI
/translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
CDS 5406..5426
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5435..5452
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(5475..5492)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 5481..5705
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind complement(5776..5792)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5776..5792)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(5789..5811)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 5800..5816
/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(5824..5854)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 5869..5890
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(6007..6024)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(6178..6766)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(6258..6277)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(6937..7797)
/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 7560..7579
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(7798..7902)
/gene="bla"
/label=AmpR promoter
primer_bind complement(7981..8000)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 gcgttgacat tgattattga ctagttatta atagtaatca attacggggt cattagttca
61 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
121 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
181 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
241 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
301 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
361 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
421 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
481 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
541 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa
601 ccgtcagatc cgctagagat ccgcggccgc taatacgact cactataggg agagccgcca
661 ccatgcccaa gaagaagagg aaagtcatga gctcagagac tggcccagtg gctgtggacc
721 ccacattgag acggcggatc gagccccatg agtttgaggt attcttcgat ccgagagagc
781 tccgcaagga gacctgcctg ctttacgaaa ttaattgggg gggccggcac tccatttggc
841 gacatacatc acagaacact aacaagcacg tcgaagtcaa cttcatcgag aagttcacga
901 cagaaagata tttctgtccg aacacaaggt gcagcattac ctggtttctc agctacagcc
961 catgcggcga atgtagtagg gccatcactg aattcctgtc aaggtatccc cacgtcactc
1021 tgtttattta catcgcaagg ctgtaccacc acgctgaccc cgaaaatcga caaggcctgg
1081 aggatttgat ctcttcaggt gtgactatcc aaattatgac tgagcaggag tcaggatact
1141 gctggagaaa ctttgtgaat tatagcccga gtaatgaagc ccactggcct aggtatcccc
1201 atctgtgggt acgactgtac gttcttgaac tgtactgcat catactgggc ctgcctcctt
1261 gtctcaacat tctgagaagg aagcagccac agctgacatt ctttaccatc gctcttcagt
1321 cttgtcatta ccagcgactg cccccacaca ttctctgggc caccgggttg aaaagcggca
1381 gcgagactcc cgggacctca gagtccgcca cacccgaaag tatgtcaaag ctggagaaat
1441 tcaccaactg ttatagcctg tctaagaccc tgcgcttcaa ggcaatccca gtgggcaaga
1501 cacaagagaa cattgacaac aaacggctcc tggtggagga tgagaagagg gctgaagatt
1561 acaagggcgt taagaagctg ctggataggt actatctgtc attcatcaac gatgtcctcc
1621 acagtatcaa gctgaagaat ctgaacaatt acatttctct gttccggaag aagacacgga
1681 ccgagaagga gaacaaagag ctggagaatc tggagatcaa cctgaggaaa gaaatagcta
1741 aggctttcaa agggaacgag ggttacaagt ccctgttcaa gaaagacatt atcgagacta
1801 ttctgcctga gttcctggac gataaagatg agatcgccct cgtcaattcc ttcaatgggt
1861 ttaccacagc ctttaccggc ttcttcgaca atagagagaa tatgttctct gaagaggcca
1921 aatccactag catcgccttt cgctgcataa acgagaacct gactaggtac atcagcaata
1981 tggacatctt tgagaaagtc gatgccatat tcgacaaaca tgaggtgcag gagattaagg
2041 agaagatcct gaactcagat tacgatgtcg aagatttctt cgagggagag ttcttcaact
2101 tcgtgctcac acaagagggc attgatgtgt acaatgcaat cattggaggg ttcgtgacag
2161 agagtggcga gaagataaag ggcctgaacg agtatatcaa cctctacaac cagaaaacca
2221 agcagaaact gcctaagttc aagccactgt acaaacaagt gctctcagat agggaaagcc
2281 tgagcttcta cggtgaaggg tatacatcag atgaagaagt gctcgaagtg ttccgcaaca
2341 ccctcaataa gaacagtgaa atcttctctt caatcaagaa gctggagaaa ctgttcaaga
2401 atttcgatga gtactcctct gccggaatct ttgtgaagaa tggccctgca atatccacta
2461 ttagcaaaga catctttggc gagtggaacg ttatcaggga taagtggaat gccgagtacg
2521 atgatattca tctcaagaag aaagccgtgg ttacagagaa atacgaggat gatagacgca
2581 agagctttaa gaagattggt agcttctctc tcgaacagct gcaggagtac gccgacgctg
2641 acctgtcagt cgtggagaaa ctcaaggaga tcataatcca gaaggtggat gaaatctaca
2701 aagtgtatgg aagctctgag aaactcttcg atgcagactt tgttctggag aagagtctga
2761 agaagaacga cgcagtggtt gctatcatga aggacctgct ggattctgtt aagtctttcg
2821 agaattacat taaggcattc tttggtgaag ggaaggagac aaatagggac gagagcttct
2881 atggcgactt tgttctggcc tacgacatcc tcctcaaggt tgaccacatc tatgacgcta
2941 tacggaatta cgttacccag aagccctata gcaaagacaa gttcaagctg tatttccaga
3001 atccacagtt tatgggtggg tgggataaag acaaagaaac agattacagg gccactatcc
3061 tgcggtacgg cagcaaatac tatctggcta tcatggataa gaagtacgcc aaatgcctcc
3121 agaagatcga caaggacgac gtgaacggta actacgagaa gatcaattac aagctcctgc
3181 caggacctaa caagatgctg cccaaggtgt tcttctccaa gaaatggatg gcctactata
3241 acccaagcga ggacattcag aagatataca agaatgggac attcaagaag ggcgatatgt
3301 tcaacctcaa cgactgccac aagctgattg atttcttcaa ggatagcatt tctcgctatc
3361 ccaagtggtc taatgcatac gatttcaact tcagcgagac tgagaagtac aaagacatcg
3421 ctggcttcta ccgggaggtg gaagagcaag gctataaggt gtcattcgaa tccgcttcta
3481 agaaggaagt ggataagctc gtggaagagg gtaagctgta catgttccag atatacaaca
3541 aagacttcag cgataagagc cacggcactc caaacctcca tactatgtat ttcaagctgc
3601 tgtttgacga gaacaaccac ggacagatta ggctgtcagg aggcgcagaa ctcttcatgc
3661 gcagagcttc actgaagaag gaggaactcg ttgtccaccc agccaatagc cctatagcca
3721 ataagaatcc agacaatcct aagaaaacca ctactctgtc ttacgatgtg tataaggata
3781 agagattctc tgaagatcag tacgaactgc acatacccat tgccattaac aagtgcccta
3841 agaacatctt caagattaac acagaggtta gagtgctcct gaaacacgac gataaccctt
3901 atgttatagg cattgctcgc ggagagagaa acctgctgta catcgtcgtg gtggacggca
3961 aaggcaacat cgtggaacag tacagtctca atgaaatcat taacaatttc aacggaatcc
4021 gcattaagac cgactaccat tctctcctcg acaagaagga gaaagaaagg ttcgaagcaa
4081 gacagaattg gacaagtata gagaatatca aagaactgaa ggctgggtac atctctcagg
4141 ttgtgcacaa gatatgtgag ctggtggaga agtacgacgc tgttatcgcc ctcgcggacc
4201 tgaatagcgg cttcaagaac tccagggtga aggtggagaa gcaggtgtat cagaagttcg
4261 agaagatgct gatcgacaag ctcaactata tggtggacaa gaaatccaat ccttgcgcta
4321 ctggtggagc cctgaagggc tatcaaatca ccaataagtt cgaatctttc aagtctatga
4381 gcacccagaa tggcttcatc ttctacatac ccgcatggct gacatccaag attgatccct
4441 ctaccggatt tgttaatctg ctcaagacta agtacacctc tattgctgac tcaaagaagt
4501 tcatatcatc atttgaccgc atcatgtacg tgccagaaga ggacctgttc gagtttgccc
4561 tggattacaa gaatttctct cggactgacg ccgactacat caagaagtgg aagctctact
4621 cttatggtaa tcggattcgc atattccgca atcccaagaa gaataacgtg ttcgattggg
4681 aggaagtttg cctcaccagc gcttacaagg agctgttcaa taagtatggg attaactacc
4741 agcagggcga cataagagcc ctgctgtgcg aacaatctga taaggcattc tattcctctt
4801 tcatggcact gatgtcactg atgctgcaaa tgcgcaattc catcaccgga agaacagacg
4861 tggcctttct gatctctcct gtcaagaact cagatggcat cttctacgat tcccgcaact
4921 atgaagcaca ggagaatgct atcctgccta agaatgccga tgcaaatgga gcctataaca
4981 tcgccagaaa ggtcctctgg gccataggac aattcaagaa agctgaagat gagaagctgg
5041 acaaggtgaa gatcgccatt tcaaacaaag agtggctcga atatgctcag acctcagtga
5101 agcatggatc acccaagaag aaacggaaag tgtctggtgg ttctactaat ctgtcagata
5161 ttattgaaaa ggagaccggt aagcaactgg ttatccagga atccatcctc atgctcccag
5221 aggaggtgga agaagtcatt gggaacaagc cggaaagcga tatactcgtg cacaccgcct
5281 acgacgagag caccgacgag aatgtcatgc ttctgactag cgacgcccct gaatacaagc
5341 cttgggctct ggtcatacag gatagcaacg gtgagaacaa gattaagatg ctctctggtg
5401 gttctcccaa gaagaagagg aaagtctaac cggtcatcat caccatcacc attgagttta
5461 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
5521 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
5581 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
5641 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
5701 tatggcttct gaggcggaaa gaaccagctg gggctcgata ccgtcgacct ctagctagag
5761 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
5821 acacaacata cgagccggaa gcataaagtg taaagcctag ggtgcctaat gagtgagcta
5881 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
5941 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
6001 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
6061 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
6121 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
6181 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
6241 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
6301 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
6361 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
6421 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
6481 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
6541 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
6601 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
6661 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
6721 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
6781 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
6841 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
6901 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
6961 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
7021 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
7081 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
7141 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
7201 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
7261 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
7321 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
7381 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
7441 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
7501 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
7561 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
7621 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
7681 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
7741 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
7801 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
7861 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
7921 gccacctgac gtcgacggat cgggagatcg atctcccgat cccctagggt cgactctcag
7981 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga
8041 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
8101 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
8161 atatac
//
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