| 质粒名称: | ERroGFP_iE_pCDNA3 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 47954 |
| 存储实验室: | David Ron |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA3 derivative |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 5370 bp |
| 完整质粒大小: | 6256 bp |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 新霉素（G418） |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | ERroGFP |
| 物种来源: | -- |
| 插入大小: | 886 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | AgeI(未破坏) |
| 3' 克隆位点: | NotI (未破坏) |
| 5' 测序引物 | CMV-F |
| 3' 测序引物 | SV40pA-R |
|
文献引用方法 |
|
| 材料和方法部分: | ERroGFP_iE_pCDNA3 was a gift from David Ron (Addgene plasmid # 47954 ; http://n2t.net/addgene:47954 ; RRID:Addgene_47954) |
| 参考文献部分: | Lifetime imaging of a fluorescent protein sensor reveals surprising stability of ER thiol redox. Avezov E, Cross BC, Kaminski Schierle GS, Winters M, Harding HP, Melo EP, Kaminski CF, Ron D. J Cell Biol. 2013 Apr 15;201(2):337-49. doi: 10.1083/jcb.201211155. 10.1083/jcb.201211155 PubMed 23589496 |
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 ERroGFP_iE_pCDNA3 6256 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Mammalian expression vector encoding for the ER targeted and FLAGM1
tagged redox probe roGFPiE (roGFP C147, E147b, C204) .
ACCESSION .
VERSION .
KEYWORDS ERroGFP_iE_pCDNA3
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6256)
AUTHORS Avezov E, Cross BC, Kaminski Schierle GS, Winters M, Harding HP,
Melo EP, Kaminski CF, Ron D
TITLE Lifetime imaging of a fluorescent protein sensor reveals surprising
stability of ER thiol redox.
JOURNAL J Cell Biol. 2013 Apr 15;201(2):337-49. doi: 10.1083/jcb.201211155.
PUBMED 23589496
REFERENCE 2 (bases 1 to 6256)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6256
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(44..63)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 235..614
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 615..818
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 769..789
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 815..839
/label=LNCX
/note="Human CMV promoter, forward primer"
CDS 949..972
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
primer_bind complement(1006..1034)
/label=GFP-R
/note="GFP, reverse primer. Does NOT anneal to EGFP"
primer_bind 1632..1652
/label=GFP-F
/note="GFP, forward primer. Does NOT anneal to EGFP"
promoter complement(1808..1826)
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind complement(1809..1826)
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind complement(1846..1863)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 1852..2076
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 2122..2550
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(2209..2228)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 2419..2440
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(2559..2579)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 2564..2894
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 2745..2880
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 2807..2826
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 2961..3755
/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
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(3015..3034)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 3625..3644
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 3929..4050
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(3966..3985)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 4020..4039
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(4099..4115)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(4099..4115)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(4112..4134)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 4123..4139
/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(4147..4177)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4192..4213
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(4330..4347)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(4501..5089)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(4581..4600)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(5260..6120)
/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 5883..5902
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(6121..6225)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
781 gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagatcg
841 cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgactc tagaggatcc
901 gccatgagcg ccctgctgat cctggctctc gtgggcgctg ccgtggccga ctacaaggac
961 gacgacgaca agtccggaag taaaggagaa gaacttttca ctggagttgt cccaattctt
1021 gttgaattag atggtgatgt taatgggcac aaattttctg tcagtggaga gggtgaaggt
1081 gatgcaacat acggaaaact tacccttaaa tttatttcca ctactggaaa actacctgtt
1141 ccatggccaa cacttgtcac tactctgagt tatggtgttc aatgcttttc aagataccca
1201 gatcatatga aacggcatga ctttttcaag agtgccatgc ccgaaggtta tgtacaggaa
1261 agaactataa gcttcaaaga tgacgggaac tacaagacac gtgctgaagt caagtttgaa
1321 ggtgataccc ttgttaatag aatcgagtta aaaggtattg attttaaaga agatggaaac
1381 attcttggac acaaattgga atacaactat aactgcgagt ccaatgtata catcatggca
1441 gacaaacaaa agaatggaat caaagcgaac ttcaaaacca gacacaacat tgaagatgga
1501 agcgttcaac tagcagacca ttatcaacaa aatactccaa ttggcgatgg ccctgtcctt
1561 ttaccagaca accattacct gtccacatgc tctgcccttt cgaaagatcc caacgaaaag
1621 agagaccaca tggtccttct tgagtttgta acagctgctg ggattacaca tggcatggat
1681 gaactataca aagatgagct ataaggtacc gagctcggat ccactagtaa cggccgccag
1741 tgtgctggaa ttctgcagat atccatcaca ctggcggccg ctcgagcatg catctagagg
1801 gccctattct atagtgtcac ctaaatgcta gagctcgctg atcagcctcg actgtgcctt
1861 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
1921 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
1981 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
2041 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
2101 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
2161 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
2221 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcggggca
2281 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
2341 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
2401 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
2461 cggtctattc ttttgattta taagggattt tggggatttc ggcctattgg ttaaaaaatg
2521 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg
2581 tggaaagtcc ccaggctccc caggcaggca gaagtatgca aagcatgcat ctcaattagt
2641 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
2701 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
2761 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
2821 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
2881 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga
2941 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
3001 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
3061 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
3121 ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg
3181 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
3241 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
3301 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
3361 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
3421 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
3481 tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
3541 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
3601 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
3661 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
3721 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
3781 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
3841 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
3901 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
3961 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
4021 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
4081 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
4141 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
4201 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
4261 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
4321 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
4381 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
4441 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
4501 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
4561 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
4621 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
4681 cgtggcgctt tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
4741 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
4801 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
4861 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
4921 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
4981 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
5041 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
5101 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
5161 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
5221 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
5281 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
5341 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
5401 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
5461 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
5521 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
5581 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
5641 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
5701 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
5761 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
5821 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
5881 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
5941 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
6001 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
6061 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
6121 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
6181 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
6241 agtgccacct gacgtc
//
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