| 质粒名称: | GFP-p53 |
|---|---|
| 出品公司: | addgene |
| 目录编号: | 12091 |
| 存储实验室: | Tyler Jacks |
|
载体骨架基本信息 |
|
| 载体名称: | pEGFP-N1 |
| 载体出品公司: | Clontech |
| 空载体大小: | 4730 bp |
| 完整质粒大小: | 5894 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: | p53 |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 951 bp |
| 基因突变: | 72R variant of p53 |
| GenBank编号 | AAD28628.1 |
| 启动子: | CMV |
| 标签/融合蛋白: | GFP (C terminal on backbone)） |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | PstI(未破坏) |
| 3’克隆位点: | BamHI(未破坏) |
| 5’测序引物 | CMV-F |
| 3’测序引物 | EGFP-N |
|
文献引用方法 |
|
| 材料和方法部分: | GFP-p53 was a gift from Tyler Jacks (Addgene plasmid # 12091 ; http://n2t.net/addgene:12091 ; RRID:Addgene_12091) |
| 参考文献部分: | An intact HDM2 RING-finger domain is required for nuclear exclusion of p53. Boyd SD, Tsai KY, Jacks T. Nat Cell Biol. 2000 Sep . 2(9):563-8. 10.1038/35023500 PubMed 10980695 |
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 GFP-p53 5894 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS GFP-p53
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5894)
AUTHORS Boyd SD, Tsai KY, Jacks T
TITLE An intact HDM2 RING-finger domain is required for nuclear exclusion
of p53.
JOURNAL Nat Cell Biol. 2000 Sep . 2(9):563-8.
PUBMED 10980695
REFERENCE 2 (bases 1 to 5894)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..5894
/organism="synthetic DNA construct"
/mol_type="other DNA"
rep_origin 1..589
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 490..509
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
enhancer 764..1067
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 1068..1271
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 1222..1242
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
CDS 1343..2521
/codon_start=1
/gene="Homo sapiens TP53"
/product="human tumor suppressor protein that is often
mutated in cancers"
/label=p53
/note="responds to cellular stresses to regulate the
expression of target genes"
/translation="MEEPQSDPSVEPPLSQETFSDLWKLLPENNVLSPLPSQAMDDLML
SPDDIEQWFTEDPGPDEAPRMPEAAPRVAPAPAAPTPAAPAPAPSWPLSSSVPSQKTYQ
GSYGFRLGFLHSGTAKSVTCTYSPALNKMFCQLAKTCPVQLWVDSTPPPGTRVRAMAIY
KQSQHMTEVVRRCPHHERCSDSDGLAPPQHLIRVEGNLRVEYLDDRNTFRHSVVVPYEP
PEVGSDCTTIHYNYMCNSSCMGGMNRRPILTIITLEDSSGNLLGRNSFEVRVCACPGRD
RRTEEENLRKKGEPHHELPPGSTKRALPNNTSSSPQPKKKPLDGEYFTLQIRGRERFEM
FRELNEALELKDAQAGKEPGGSRAHSSHLKSKKGQSTSRHKKLMFKTEGPDSD"
regulatory 2537..2546
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2543..3262
/codon_start=1
/product="the original enhanced GFP (Yang et al., 1996)"
/label=EGFP
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
CDS 2543..3259
/codon_start=1
/product="enhanced GFP"
/label=EGFP
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
primer_bind complement(2588..2609)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(2849..2868)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
primer_bind 3196..3217
/label=EGFP-C
/note="EGFP, forward primer"
polyA_signal 3385..3506
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(3422..3441)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 3476..3495
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
rep_origin complement(3513..3968)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(3650..3671)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 3862..3881
/label=F1ori-R
/note="F1 origin, reverse primer"
promoter 3995..4099
/gene="bla"
/label=AmpR promoter
promoter 4101..4458
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
primer_bind complement(4124..4144)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 4309..4444
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 4371..4390
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 4493..5287
/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
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(4547..4566)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 5157..5176
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
primer_bind complement(5475..5494)
/label=TK-pA-R
/note="Thymidine kinase polyA, reverse primer"
polyA_signal 5519..5566
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
661 gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat
721 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
781 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
841 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
901 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
961 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
1021 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
1081 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
1141 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
1201 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
1261 taagcagagc tggtttagtg aaccgtcaga tccgctagcg ctaccggact cagatctcga
1321 gctcaagctt cgaattctgc agatggagga gccgcagtca gatcctagcg tcgagccccc
1381 tctgagtcag gaaacatttt cagacctatg gaaactactt cctgaaaaca acgttctgtc
1441 ccccttgccg tcccaagcaa tggatgattt gatgctgtcc ccggacgata ttgaacaatg
1501 gttcactgaa gacccaggtc cagatgaagc tcccagaatg ccagaggctg ctccccgcgt
1561 ggcccctgca ccagcagctc ctacaccggc ggcccctgca ccagccccct cctggcccct
1621 gtcatcttct gtcccttccc agaaaaccta ccagggcagc tacggtttcc gtctgggctt
1681 cttgcattct gggacagcca agtctgtgac ttgcacgtac tcccctgccc tcaacaagat
1741 gttttgccaa ctggccaaga cctgccctgt gcagctgtgg gttgattcca cacccccgcc
1801 cggcacccgc gtccgcgcca tggccatcta caagcagtca cagcacatga cggaggttgt
1861 gaggcgctgc ccccaccatg agcgctgctc agatagcgat ggtctggccc ctcctcagca
1921 tcttatccga gtggaaggaa atttgcgtgt ggagtatttg gatgacagaa acacttttcg
1981 acatagtgtg gtggtgccct atgagccgcc tgaggttggc tctgactgta ccaccatcca
2041 ctacaactac atgtgtaaca gttcctgcat gggcggcatg aaccggaggc ccatcctcac
2101 catcatcaca ctggaagact ccagtggtaa tctactggga cggaacagct ttgaggtgcg
2161 tgtttgtgcc tgtcctggga gagaccggcg cacagaggaa gagaatctcc gcaagaaagg
2221 ggagcctcac cacgagctgc ccccagggag cactaagcga gcactgccca acaacaccag
2281 ctcctctccc cagccaaaga agaaaccact ggatggagaa tatttcaccc ttcagatccg
2341 tgggcgtgag cgcttcgaga tgttccgaga gctgaatgag gccttggaac tcaaggatgc
2401 ccaggctggg aaggagccag gggggagcag ggctcactcc agccacctga agtccaaaaa
2461 gggtcagtct acctcccgcc ataaaaaact catgttcaag acagaagggc ctgactcaga
2521 ccgggatcca ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt
2581 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
2641 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
2701 gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag
2761 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
2821 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
2881 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
2941 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
3001 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
3061 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
3121 cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa
3181 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
3241 catggacgag ctgtacaagt aaagcggccg cgactctaga tcataatcag ccataccaca
3301 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
3361 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
3421 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
3481 ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa
3541 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
3601 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
3661 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
3721 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
3781 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
3841 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
3901 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
3961 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
4021 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
4081 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
4141 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
4201 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
4261 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
4321 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
4381 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
4441 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
4501 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
4561 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
4621 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
4681 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
4741 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
4801 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
4861 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
4921 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
4981 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
5041 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
5101 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
5161 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
5221 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
5281 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
5341 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
5401 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct
5461 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
5521 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
5581 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
5641 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
5701 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
5761 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
5821 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
5881 atcaaaggat cttc
//
