| 质粒名称: | pCMV-dCpf1-BE-YE |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 107686 |
| 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: | dLbCpf1-BE-YE / pCMV-SV40NLS-APOBEC1(W90Y/R126E)-XTEN-dLbCpf1(D832A/E1006A/D1125A)-SV40NLS-SGGS-UGI-SV40NLS |
| 物种来源: | R. norvegicus (rat); Lachnospiraceae bacterium Bacteriophage PBS2 |
| 插入大小: | 4764 bp |
| 基因突变: | APOBEC1(W90Y/R126E), dLbCpf1(D832A/E925A/D1148A) |
| 启动子: | CMV |
| 克隆方法: | Gibson Cloning |
| 5’克隆位点: | AgeI(未破坏) |
| 3’克隆位点: | NotI (未破坏) |
| 5’测序引物 | CMV |
| 3’测序引物 | BGH poly reverse |
|
文献引用方法 |
|
| 材料和方法部分: | pCMV-dCpf1-BE-YE was a gift from Jia Chen (Addgene plasmid # 107686 ; http://n2t.net/addgene:107686 ; RRID:Addgene_107686) |
| 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-YE 8166 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Expresses dLbCpf1-BE-YE in mammalian cells.
ACCESSION .
VERSION .
KEYWORDS pCMV-dCpf1-BE-YE
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 16..395
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 396..599
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 550..570
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 641..660
/label=T7
/note="T7 promoter, forward primer"
promoter 641..659
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 676..696
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5122..5142
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5155..5403
/codon_start=1
/product="uracil-DNA glycosylase inhibitor from a Bacillus
subtilis bacteriophage (Mol et al., 1995)"
/label=UGI
/translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
CDS 5416..5436
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5445..5462
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(5485..5502)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 5491..5715
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind complement(5786..5802)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5786..5802)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(5799..5821)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 5810..5826
/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(5834..5864)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 5879..5900
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(6017..6034)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(6188..6776)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(6268..6287)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(6947..7807)
/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 7570..7589
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(7808..7912)
/gene="bla"
/label=AmpR promoter
primer_bind complement(7991..8010)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca attacggggt
61 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
121 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
181 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
241 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
301 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
361 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
421 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
481 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
541 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctg
601 gtttagtgaa ccgtcagatc cgctagagat ccgcggccgc taatacgact cactataggg
661 agagccgcca ccatgcccaa gaagaagagg aaagtcatga gctcagagac tggcccagtg
721 gctgtggacc ccacattgag acggcggatc gagccccatg agtttgaggt attcttcgat
781 ccgagagagc tccgcaagga gacctgcctg ctttacgaaa ttaattgggg gggccggcac
841 tccatttggc gacatacatc acagaacact aacaagcacg tcgaagtcaa cttcatcgag
901 aagttcacga cagaaagata tttctgtccg aacacaaggt gcagcattac ctggtttctc
961 agctacagcc catgcggcga atgtagtagg gccatcactg aattcctgtc aaggtatccc
1021 cacgtcactc tgtttattta catcgcaagg ctgtaccacc acgctgaccc cgaaaatcga
1081 caaggcctgc gggatttgat ctcttcaggt gtgactatcc aaattatgac tgagcaggag
1141 tcaggatact gctggagaaa ctttgtgaat tatagcccga gtaatgaagc ccactggcct
1201 aggtatcccc atctgtgggt acgactgtac gttcttgaac tgtactgcat catactgggc
1261 ctgcctcctt gtctcaacat tctgagaagg aagcagccac agctgacatt ctttaccatc
1321 gctcttcagt cttgtcatta ccagcgactg cccccacaca ttctctgggc caccgggttg
1381 aaaagcggca gcgagactcc cgggacctca gagtccgcca cacccgaaag tatgtcaaag
1441 ctggagaaat tcaccaactg ttatagcctg tctaagaccc tgcgcttcaa ggcaatccca
1501 gtgggcaaga cacaagagaa cattgacaac aaacggctcc tggtggagga tgagaagagg
1561 gctgaagatt acaagggcgt taagaagctg ctggataggt actatctgtc attcatcaac
1621 gatgtcctcc acagtatcaa gctgaagaat ctgaacaatt acatttctct gttccggaag
1681 aagacacgga ccgagaagga gaacaaagag ctggagaatc tggagatcaa cctgaggaaa
1741 gaaatagcta aggctttcaa agggaacgag ggttacaagt ccctgttcaa gaaagacatt
1801 atcgagacta ttctgcctga gttcctggac gataaagatg agatcgccct cgtcaattcc
1861 ttcaatgggt ttaccacagc ctttaccggc ttcttcgaca atagagagaa tatgttctct
1921 gaagaggcca aatccactag catcgccttt cgctgcataa acgagaacct gactaggtac
1981 atcagcaata tggacatctt tgagaaagtc gatgccatat tcgacaaaca tgaggtgcag
2041 gagattaagg agaagatcct gaactcagat tacgatgtcg aagatttctt cgagggagag
2101 ttcttcaact tcgtgctcac acaagagggc attgatgtgt acaatgcaat cattggaggg
2161 ttcgtgacag agagtggcga gaagataaag ggcctgaacg agtatatcaa cctctacaac
2221 cagaaaacca agcagaaact gcctaagttc aagccactgt acaaacaagt gctctcagat
2281 agggaaagcc tgagcttcta cggtgaaggg tatacatcag atgaagaagt gctcgaagtg
2341 ttccgcaaca ccctcaataa gaacagtgaa atcttctctt caatcaagaa gctggagaaa
2401 ctgttcaaga atttcgatga gtactcctct gccggaatct ttgtgaagaa tggccctgca
2461 atatccacta ttagcaaaga catctttggc gagtggaacg ttatcaggga taagtggaat
2521 gccgagtacg atgatattca tctcaagaag aaagccgtgg ttacagagaa atacgaggat
2581 gatagacgca agagctttaa gaagattggt agcttctctc tcgaacagct gcaggagtac
2641 gccgacgctg acctgtcagt cgtggagaaa ctcaaggaga tcataatcca gaaggtggat
2701 gaaatctaca aagtgtatgg aagctctgag aaactcttcg atgcagactt tgttctggag
2761 aagagtctga agaagaacga cgcagtggtt gctatcatga aggacctgct ggattctgtt
2821 aagtctttcg agaattacat taaggcattc tttggtgaag ggaaggagac aaatagggac
2881 gagagcttct atggcgactt tgttctggcc tacgacatcc tcctcaaggt tgaccacatc
2941 tatgacgcta tacggaatta cgttacccag aagccctata gcaaagacaa gttcaagctg
3001 tatttccaga atccacagtt tatgggtggg tgggataaag acaaagaaac agattacagg
3061 gccactatcc tgcggtacgg cagcaaatac tatctggcta tcatggataa gaagtacgcc
3121 aaatgcctcc agaagatcga caaggacgac gtgaacggta actacgagaa gatcaattac
3181 aagctcctgc caggacctaa caagatgctg cccaaggtgt tcttctccaa gaaatggatg
3241 gcctactata acccaagcga ggacattcag aagatataca agaatgggac attcaagaag
3301 ggcgatatgt tcaacctcaa cgactgccac aagctgattg atttcttcaa ggatagcatt
3361 tctcgctatc ccaagtggtc taatgcatac gatttcaact tcagcgagac tgagaagtac
3421 aaagacatcg ctggcttcta ccgggaggtg gaagagcaag gctataaggt gtcattcgaa
3481 tccgcttcta agaaggaagt ggataagctc gtggaagagg gtaagctgta catgttccag
3541 atatacaaca aagacttcag cgataagagc cacggcactc caaacctcca tactatgtat
3601 ttcaagctgc tgtttgacga gaacaaccac ggacagatta ggctgtcagg aggcgcagaa
3661 ctcttcatgc gcagagcttc actgaagaag gaggaactcg ttgtccaccc agccaatagc
3721 cctatagcca ataagaatcc agacaatcct aagaaaacca ctactctgtc ttacgatgtg
3781 tataaggata agagattctc tgaagatcag tacgaactgc acatacccat tgccattaac
3841 aagtgcccta agaacatctt caagattaac acagaggtta gagtgctcct gaaacacgac
3901 gataaccctt atgttatagg cattgctcgc ggagagagaa acctgctgta catcgtcgtg
3961 gtggacggca aaggcaacat cgtggaacag tacagtctca atgaaatcat taacaatttc
4021 aacggaatcc gcattaagac cgactaccat tctctcctcg acaagaagga gaaagaaagg
4081 ttcgaagcaa gacagaattg gacaagtata gagaatatca aagaactgaa ggctgggtac
4141 atctctcagg ttgtgcacaa gatatgtgag ctggtggaga agtacgacgc tgttatcgcc
4201 ctcgcggacc tgaatagcgg cttcaagaac tccagggtga aggtggagaa gcaggtgtat
4261 cagaagttcg agaagatgct gatcgacaag ctcaactata tggtggacaa gaaatccaat
4321 ccttgcgcta ctggtggagc cctgaagggc tatcaaatca ccaataagtt cgaatctttc
4381 aagtctatga gcacccagaa tggcttcatc ttctacatac ccgcatggct gacatccaag
4441 attgatccct ctaccggatt tgttaatctg ctcaagacta agtacacctc tattgctgac
4501 tcaaagaagt tcatatcatc atttgaccgc atcatgtacg tgccagaaga ggacctgttc
4561 gagtttgccc tggattacaa gaatttctct cggactgacg ccgactacat caagaagtgg
4621 aagctctact cttatggtaa tcggattcgc atattccgca atcccaagaa gaataacgtg
4681 ttcgattggg aggaagtttg cctcaccagc gcttacaagg agctgttcaa taagtatggg
4741 attaactacc agcagggcga cataagagcc ctgctgtgcg aacaatctga taaggcattc
4801 tattcctctt tcatggcact gatgtcactg atgctgcaaa tgcgcaattc catcaccgga
4861 agaacagacg tggcctttct gatctctcct gtcaagaact cagatggcat cttctacgat
4921 tcccgcaact atgaagcaca ggagaatgct atcctgccta agaatgccga tgcaaatgga
4981 gcctataaca tcgccagaaa ggtcctctgg gccataggac aattcaagaa agctgaagat
5041 gagaagctgg acaaggtgaa gatcgccatt tcaaacaaag agtggctcga atatgctcag
5101 acctcagtga agcatggatc acccaagaag aaacggaaag tgtctggtgg ttctactaat
5161 ctgtcagata ttattgaaaa ggagaccggt aagcaactgg ttatccagga atccatcctc
5221 atgctcccag aggaggtgga agaagtcatt gggaacaagc cggaaagcga tatactcgtg
5281 cacaccgcct acgacgagag caccgacgag aatgtcatgc ttctgactag cgacgcccct
5341 gaatacaagc cttgggctct ggtcatacag gatagcaacg gtgagaacaa gattaagatg
5401 ctctctggtg gttctcccaa gaagaagagg aaagtctaac cggtcatcat caccatcacc
5461 attgagttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
5521 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
5581 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
5641 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
5701 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctcgata ccgtcgacct
5761 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
5821 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctag ggtgcctaat
5881 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
5941 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
6001 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
6061 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
6121 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
6181 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
6241 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
6301 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
6361 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
6421 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
6481 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
6541 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
6601 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
6661 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
6721 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
6781 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
6841 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
6901 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
6961 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
7021 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
7081 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
7141 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
7201 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
7261 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
7321 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
7381 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
7441 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
7501 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
7561 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
7621 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
7681 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
7741 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
7801 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
7861 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
7921 cccgaaaagt gccacctgac gtcgacggat cgggagatcg atctcccgat cccctagggt
7981 cgactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
8041 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
8101 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
8161 tacggg
//
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