| 质粒名称: | GFP-Nucleolin |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 28176 |
| 存储实验室: | Michael Kastan |
|
载体骨架基本信息 |
|
| 载体名称: | pEGFPC1 |
| 载体出品公司: | Clontech |
| 空载体大小: | 4700 bp |
| 完整质粒大小: | 6987 bp |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 新霉素（G418） |
| 载体抗性: | Kanamycin |
| 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: | Nucleolin |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 2100bp |
| 标签/融合蛋白: | GFP (N terminal on backbone) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | KpnI(未破坏) |
| 3’克隆位点: | BamHI (未破坏) |
| 5’测序引物 | EGFP-C |
| 3’测序引物 | SV40-pA-R |
|
文献引用方法 |
|
| 材料和方法部分: | GFP-Nucleolin was a gift from Michael Kastan (Addgene plasmid # 28176 ; http://n2t.net/addgene:28176 ; RRID:Addgene_28176) |
| 参考文献部分: | Regulation of p53 translation and induction after DNA damage by ribosomal protein L26 and nucleolin. Takagi M, Absalon MJ, McLure KG, Kastan MB. Cell. 2005 Oct 7. 123(1):49-63. 10.1016/j.cell.2005.07.034 PubMed 16213212 |
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-Nucleolin 6987 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS GFP-Nucleolin
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6987)
AUTHORS Takagi M, Absalon MJ, McLure KG, Kastan MB
TITLE Regulation of p53 translation and induction after DNA damage by
ribosomal protein L26 and nucleolin.
JOURNAL Cell. 2005 Oct 7. 123(1):49-63.
PUBMED 16213212
REFERENCE 2 (bases 1 to 6987)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..6987
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 68..371
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 372..575
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 526..546
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
regulatory 614..623
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 620..1336
/codon_start=1
/product="enhanced GFP"
/label=EGFP
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
CDS 620..1336
/codon_start=1
/product="the original enhanced GFP (Yang et al., 1996)"
/label=EGFP
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
primer_bind complement(665..686)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(926..945)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
primer_bind 1273..1294
/label=EGFP-C
/note="EGFP, forward primer"
protein_bind 1931..1942
/label=ZF binding site
/bound_moiety="CCR5TC zinc-finger domain"
/note="target sequence for the CCR5TC zinc-finger domain
from the CCR5-224 (+) zinc-finger nuclease (Perez et al.,
2008; Gross et al., 2013)"
protein_bind 1949..1960
/label=ZF binding site
/bound_moiety="CCR5TC zinc-finger domain"
/note="target sequence for the CCR5TC zinc-finger domain
from the CCR5-224 (+) zinc-finger nuclease (Perez et al.,
2008; Gross et al., 2013)"
CDS 3008..3019
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
polyA_signal 3523..3657
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
polyA_signal 3782..3903
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin complement(3910..4365)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4047..4068)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 4259..4278
/label=F1ori-R
/note="F1 origin, reverse primer"
promoter 4392..4496
/gene="bla"
/label=AmpR promoter
promoter 4498..4855
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
primer_bind complement(4521..4541)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 4706..4841
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 4768..4787
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 4890..5684
/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(4944..4963)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 5554..5573
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
primer_bind complement(5872..5891)
/label=TK-pA-R
/note="Thymidine kinase polyA, reverse primer"
polyA_signal 5916..5963
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
rep_origin 6292..6880
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6781..6800
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
601 agcgctaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
661 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
721 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
781 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
841 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
901 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
961 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
1021 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
1081 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
1141 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
1201 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
1261 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
1321 ggacgagctg tacaagtact cagatctcga gctcaagctt ggtaccatgg tgaagctcgc
1381 gaaggcaggt aaaaatcaag gtgaccccaa gaaaatggct cctcctccaa aggaggtaga
1441 agaagatagt gaagatgagg aaatgtcaga agatgaagaa gatgatagca gtggagaaga
1501 ggtcgtcata cctcagaaga aaggcaagaa ggctgctgca acctcagcaa agaaggtggt
1561 cgtttcccca acaaaaaagg ttgcagttgc cacaccagcc aagaaagcag ctgtcactcc
1621 aggcaaaaag gcagcagcaa cacctgccaa gaagacagtt acaccagcca aagcagttac
1681 cacacctggc aagaagggag ccacaccagg caaagcattg gtagcaactc ctggtaagaa
1741 gggtgctgcc atcccagcca ggggggcaaa gaatggcaag aatgccgagg aggaagacag
1801 tgatgaagag gaggatgatg acagtgagga ggatgaggag gatgacgagg acgaggatga
1861 ggatgaagat gaaattgaac cagcagcgat gaaagcagca gctgctgccc ctgcctcaga
1921 ggatgaggac gatgaggatg acgaagatga tgaggatgac gatgacgatg aggaagatga
1981 ctctgaagaa gaagctatgg agactacacc agccaaagga aagaaagctg caaaagttgt
2041 tcctgtgaaa gccaagaacg tggctgagga tgaagatgaa gaagaggatg atgaggacga
2101 ggatgacgac gacgacgaag atgatgaaga tgatgatgat gaagatgatg aggaggagga
2161 agaagaggag gaggaagagc ctgtcaaaga agcacctgga aaacgaaaga aggaaatggc
2221 caaacagaaa gcagctcctg aagccaagaa acagaaagtg gaaggcacag aaccgactac
2281 ggctttcaat ctctttgttg gaaacctaaa ctttaacaaa tctgctcctg aattaaaaac
2341 tggtatcagc gatgtttttg ctaaaaatga tcttgctgtt gtggatgtca gaattggtat
2401 gactaggaaa tttggttatg tggattttga atctgctgaa gacctggaga aagcgttgga
2461 actcactggt ttgaaagtct ttggcaatga aattaaacta gagaaaccaa aaggaaaaga
2521 cagtaagaaa gagcgagatg cgagaacact tttggctaaa aatctccctt acaaagtcac
2581 tcaggatgaa ttgaaagaag tgtttgaaga tgctgcggag atcagattag tcagcaagga
2641 tgggaaaagt aaagggattg cttatattga atttaagaca gaagctgatg cagagaaaac
2701 ctttgaagaa aagcagggaa cagagatcga tgggcgatct atttccctgt actatactgg
2761 agagaaaggt caaaatcaag actatagagg tggaaagaat agcacttgga gtggtgaatc
2821 aaaaactctg gttttaagca acctctccta cagtgcaaca gaagaaactc ttcaggaagt
2881 atttgagaaa gcaactttta tcaaagtacc ccagaaccaa aatggcaaat ctaaagggta
2941 tgcatttata gagtttgctt cattcgaaga cgctaaagaa gctttaaatt cctgtaataa
3001 aagggaaatt gagggcagag caatcaggct ggagttgcaa ggacccaggg gatcacctaa
3061 tgccagaagc cagccatcca aaactctgtt tgtcaaaggc ctgtctgagg ataccactga
3121 agagacatta aaggagtcat ttgacggctc cgttcgggct aggatagtta ctgaccggga
3181 aactgggtcc tccaaagggt ttggttttgt agacttcaac agtgaggagg atgccaaagc
3241 tgccaaggag gccatggaag acggtgaaat tgatggaaat aaagttacct tggactgggc
3301 caaacctaag ggtgaaggtg gcttcggggg tcgtggtgga ggcagaggcg gctttggagg
3361 acgaggtggt ggtagaggag gccgaggagg atttggtggc agaagccggg gaggctttgg
3421 agggcgagga ggcttccgag gaggcagagg aggaggaggt gaccacaagc cacaaggaaa
3481 gaagacgaag tttgaatagc tcgagcatgc atctagagga tcaacttgtt tattgcagct
3541 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
3601 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatccac
3661 cggatctaga taactgatca taatcagcca taccacattt gtagaggttt tacttgcttt
3721 aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt
3781 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
3841 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
3901 ttaacgcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
3961 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
4021 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
4081 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
4141 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
4201 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
4261 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
4321 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc
4381 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
4441 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg
4501 aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
4561 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
4621 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
4681 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
4741 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
4801 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg
4861 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
4921 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
4981 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
5041 ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg
5101 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
5161 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
5221 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
5281 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
5341 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
5401 tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
5461 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
5521 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
5581 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
5641 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
5701 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
5761 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
5821 gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg
5881 gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac
5941 gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt
6001 cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc
6061 ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc
6121 tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt
6181 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
6241 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
6301 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
6361 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
6421 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
6481 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
6541 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
6601 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
6661 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
6721 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
6781 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
6841 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
6901 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
6961 tgattctgtg gataaccgta ttaccgc
//
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