| 质粒名称: | GAG-CRErec |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 119971 |
| 存储实验室: | Philippe Mangeot, Théophile Ohlmann, Emiliano Ricci |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA3-like |
| 空载体大小: | 7530 bp |
| 完整质粒大小: | 7529 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: | GAG-nlsCRErec |
| 物种来源: | H. sapiens (human), M. musculus (mouse) |
| 插入大小: | 2691 bp |
| 启动子: | CMV |
| 标签/融合蛋白: | nls |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | Xho I(未破坏) |
| 3’克隆位点: | Kpn I (未破坏) |
| 5’测序引物 | ACTACATCCTGGTCATCATCC |
| 3’测序引物 | CCACCTTCTGATAGGCAGCC |
|
文献引用方法 |
|
| 材料和方法部分: | GAG-CRErec was a gift from Philippe Mangeot & Théophile Ohlmann & Emiliano Ricci (Addgene plasmid # 119971 ; http://n2t.net/addgene:119971 ; RRID:Addgene_119971) |
| 参考文献部分: | Genome editing in primary cells and in vivo using viral-derived Nanoblades loaded with Cas9-sgRNA ribonucleoproteins. Mangeot PE, Risson V, Fusil F, Marnef A, Laurent E, Blin J, Mournetas V, Massourides E, Sohier TJM, Corbin A, Aube F, Teixeira M, Pinset C, Schaeffer L, Legube G, Cosset FL, Verhoeyen E, Ohlmann T, Ricci EP. Nat Commun. 2019 Jan 3;10(1):45. doi: 10.1038/s41467-018-07845-z. 10.1038/s41467-018-07845-z PubMed 30604748 |
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 GAG-CRErec 7529 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Expresses GAG (FMLV) fused with CRE recombinase for the production
of VLPs loaded with CRE protein.
ACCESSION .
VERSION .
KEYWORDS GAG-CRErec
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7529)
AUTHORS Mangeot PE, Risson V, Fusil F, Marnef A, Laurent E, Blin J,
Mournetas V, Massourides E, Sohier TJM, Corbin A, Aube F, Teixeira
M, Pinset C, Schaeffer L, Legube G, Cosset FL, Verhoeyen E, Ohlmann
T, Ricci EP
TITLE Genome editing in primary cells and in vivo using viral-derived
Nanoblades loaded with Cas9-sgRNA ribonucleoproteins.
JOURNAL Nat Commun. 2019 Jan 3;10(1):45. doi: 10.1038/s41467-018-07845-z.
PUBMED 30604748
REFERENCE 2 (bases 1 to 7529)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7529
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 9..31
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 23..40
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 24..40
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 50..69
/label=T7
/note="T7 promoter, forward primer"
promoter 50..68
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
enhancer 189..568
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 569..772
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 723..743
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 769..793
/label=LNCX
/note="Human CMV promoter, forward primer"
intron 884..1456
/label=-beta-globin intron
/note="intron from rabbit beta-globin gene"
CDS 3192..3212
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 3213..4241
/codon_start=1
/product="site-specific recombinase"
/label=Cre
/note="Cre is a site-specific recombinase from
bacteriophage P1. Recombination occurs at loxP sequences."
/translation="SNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKMLL
SVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGLP
RPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAFL
GIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVER
WISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQR
YLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLED
GD"
primer_bind complement(4326..4345)
/label=Bglob-pA-R
/note="Rabbit beta-globin polyA region, reverse primer"
polyA_signal 4391..4446
/label=-beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
primer_bind complement(4445..4464)
/label=rbglobpA-R
/note="Rabbit beta-globin polyA, reverse primer. Also
called rb-glob-pA-term-R"
primer_bind complement(4804..4820)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(4804..4820)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(4817..4839)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 4828..4844
/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(4852..4882)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4897..4918
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(5035..5052)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(5206..5794)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(5286..5305)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(5965..6825)
/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 6588..6607
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(6826..6930)
/gene="bla"
/label=AmpR promoter
rep_origin complement(6956..7411)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(7093..7114)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 7305..7324
/label=F1ori-R
/note="F1 origin, reverse primer"
ORIGIN
1 agggttttcc cagtcacgac gttgtaaaac gacggccagt gagcgcgcgt aatacgactc
61 actatagggc gaattggagc tccaccgcgg tggcggccgc tctagagagc ttggcccatt
121 gcatacgttg tatccatatc ataatatgta catttatatt ggctcatgtc caacattacc
181 gccatgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
241 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
301 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
361 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
421 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
481 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
541 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
601 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
661 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
721 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctcgtttagt
781 gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata gaagacaccg
841 ggaccgatcc agcctccggt cgaccgatcc tgagaacttc agggtgagtt tggggaccct
901 tgattgttct ttctttttcg ctattgtaaa attcatgtta tatggagggg gcaaagtttt
961 cagggtgttg tttagaatgg gaagatgtcc cttgtatcac catggaccct catgataatt
1021 ttgtttcttt cactttctac tctgttgaca accattgtct cctcttattt tcttttcatt
1081 ttctgtaact ttttcgttaa actttagctt gcatttgtaa cgaattttta aattcacttt
1141 tgtttatttg tcagattgta agtactttct ctaatcactt ttttttcaag gcaatcaggg
1201 tatattatat tgtacttcag cacagtttta gagaacaatt gttataatta aatgataagg
1261 tagaatattt ctgcatataa attctggctg gcgtggaaat attcttattg gtagaaacaa
1321 ctacaccctg gtcatcatcc tgcctttctc tttatggtta caatgatata cactgtttga
1381 gatgaggata aaatactctg agtccaaacc gggcccctct gctaaccatg ttcatgcctt
1441 cttctctttc ctacagctcc tgggcaacgt gctggttgtt gtgctgtctc atcattttgg
1501 caaagaattc ctcgagggcc ggatccgaat tcctgtattt gtctgaaaac atgggccagg
1561 ctgttaccac ccccttaagt ttgactttag accactggaa ggatgtcgaa cggacagccc
1621 acaacctgtc ggtagaggtt agaaaaaggc gctgggttac attctgctct gcagaatggc
1681 caaccttcaa cgtcggatgg ccacgagacg gcacttttaa cccagacatt attacacagg
1741 ttaagatcaa ggtcttctca cctggcccac atggacatcc ggatcaggtc ccctacatcg
1801 tgacctggga agctatagca gtagaccccc ctccctgggt cagacccttc gtgcacccta
1861 aacctcccct ctctcttccc ccttcagccc cctctctccc acctgaaccc ccactctcga
1921 ccccgcccca gtcctccctc tatccggctc tcacttctcc tttaaacacc aaacctaggc
1981 ctcaagtcct tcctgatagc ggaggaccac tcattgatct actcacggag gaccctccgc
2041 cttaccggga cccagggcca ccctctcctg acgggaacgg cgatagcgga gaagtggccc
2101 ctacagaagg agcccctgac ccttccccaa tggtatcccg cctgcgggga agaaaagaac
2161 cccccgtggc ggattctact acctctcagg cgttccccct tcgcctggga gggaatggac
2221 agtatcaata ctggccattt tcctcctctg acctctataa ctggaaaaat aacaacccct
2281 ctttctccga ggacccagct aaattgacag ctttgatcga gtccgttctc cttactcatc
2341 agcccacttg ggatgactgc caacagctat tagggaccct gctgacggga gaagaaaaac
2401 agcgagtgct cctagaggcc cgaaaggcgg ttcgagggga ggacggacgc ccaactcagc
2461 tgcccaatga cattaatgat gcttttccct tggaacgtcc cgactgggac tacaacaccc
2521 aacgaggtag gaaccaccta gtccactatc gccagttgct cctagcgggt ctccaaaacg
2581 cgggcagaag ccccaccaat ttggccaagg taaaagggat aacccaggga cctaatgagt
2641 ctccctcagc ctttttagag agactcaagg aggcctatcg cagatacact ccttatgacc
2701 ctgaggaccc agggcaagaa accaatgtgg ccatgtcatt catctggcag tccgccccgg
2761 atatcgggcg aaagttagag cggttagaag atttgaagag taagacctta ggagacttag
2821 tgagggaagc tgaaaagatc tttaataaac gagaaacccc ggaagaaaga gaggaacgta
2881 ttaggagaga aacagaggaa aaggaagaac gccgtagggc agaggatgtg cagagagaga
2941 aggagaggga ccgcagaaga catagagaaa tgagtaagtt gctggctact gtcgttagcg
3001 ggcagagaca ggatagacag ggaggagagc gaaggaggcc ccaactcgac cacgaccagt
3061 gtgcctactg caaagaaaag ggacattggg ctagagattg ccccaagaag ccaagaggac
3121 cccggggacc acgaccccag gcctccctcc tgacgcgtag ttccctgtat ccagccctca
3181 cacctaccgg ccccaagaag aagaggaagg tgtccaattt actgaccgta caccaaaatt
3241 tgcctgcatt accggtcgat gcaacgagtg atgaggttcg caagaacctg atggacatgt
3301 tcagggatcg ccaggcgttt tctgagcata cctggaaaat gcttctgtcc gtttgccggt
3361 cgtgggcggc atggtgcaag ttgaataacc ggaaatggtt tcccgcagaa cctgaagatg
3421 ttcgcgatta tcttctatat cttcaggcgc gcggtctggc agtaaaaact atccagcaac
3481 atttgggcca gctaaacatg cttcatcgtc ggtccgggct gccacgacca agtgacagca
3541 atgctgtttc actggttatg cggcggatcc gaaaagaaaa cgttgatgcc ggtgaacgtg
3601 caaaacaggc tctagcgttc gaacgcactg atttcgacca ggttcgttca ctcatggaaa
3661 atagcgatcg ctgccaggat atacgtaatc tggcatttct ggggattgct tataacaccc
3721 tgttacgtat agccgaaatt gccaggatca gggttaaaga tatctcacgt actgacggtg
3781 ggagaatgtt aatccatatt ggcagaacga aaacgctggt tagcaccgca ggtgtagaga
3841 aggcacttag cctgggggta actaaactgg tcgagcgatg gatttccgtc tctggtgtag
3901 ctgatgatcc gaataactac ctgttttgcc gggtcagaaa aaatggtgtt gccgcgccat
3961 ctgccaccag ccagctatca actcgcgccc tggaagggat ttttgaagca actcatcgat
4021 tgatttacgg cgctaaggat gactctggtc agagatacct ggcctggtct ggacacagtg
4081 cccgtgtcgg agccgcgcga gatatggccc gcgctggagt ttcaataccg gagatcatgc
4141 aagctggtgg ctggaccaat gtaaatattg tcatgaacta tatccgtaac ctggatagtg
4201 aaacaggggc aatggtgcgc ctgctggaag atggcgatta ggtacctgca tcctcaggtg
4261 caggctgcct atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac
4321 tgagatcttt ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg
4381 acttctggct aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt
4441 ctctcactcg gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg
4501 gtttagagtt tggcaacata tgcccatatg ctggctgcca tgaacaaagg ttggctataa
4561 agaggtcatc agtatatgaa acagccccct gctgtccatt ccttattcca tagaaaagcc
4621 ttgacttgag gttagatttt ttttatattt tgttttgtgt tatttttttc tttaacatcc
4681 ctaaaatttt ccttacatgt tttactagcc agatttttcc tcctctcctg actactccca
4741 gtcatagctg tccctcttct cttatggaga tccctcgacg gatcggccgc aattcgtaat
4801 catgtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
4861 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
4921 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
4981 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
5041 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
5101 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
5161 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
5221 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
5281 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
5341 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
5401 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
5461 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
5521 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
5581 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
5641 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
5701 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
5761 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
5821 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
5881 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
5941 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
6001 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
6061 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
6121 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
6181 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
6241 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
6301 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
6361 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
6421 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
6481 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
6541 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
6601 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
6661 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
6721 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
6781 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
6841 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
6901 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaaatt
6961 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
7021 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
7081 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
7141 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
7201 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
7261 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
7321 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
7381 gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct gcgcaactgt
7441 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
7501 gctgcaaggc gattaagttg ggtaacgcc
//
