| 质粒名称: | pCMV CEPIA4mt |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 58220 |
| 存储实验室: | Masamitsu Iino |
|
载体骨架基本信息 |
|
| 载体名称: | pCMV/myc/mito |
| 载体出品公司: | Invitrogen |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 4799 bp |
| 完整质粒大小: | 6404 bp |
| 载体修饰: | 重复线粒体靶向序列。 |
| 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: | CEPIA4mt,green fluorescent calcium-measuring organelle-entrapped protein indicator for the mitochondria (low affinity variant) |
| 物种来源: | Synthetic |
| 插入大小: | 1605 bp |
| 基因突变: | GCaMP2 N263Y E282V E334D M339L F395W D436E |
| GenBank编号 | -- |
| 启动子 | CMV |
| 标签/融合蛋白: | myc（插入件上的C端子） |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | XbaI (未破坏) |
| 5' 测序引物 | cgcaaatgggcggtaggcgtg |
| 3' 测序引物 | tagaaggcacagtcgagg- |
|
文献引用方法 |
|
| 材料和方法部分: | pCMV CEPIA4mt was a gift from Masamitsu Iino (Addgene plasmid # 58220 ; http://n2t.net/addgene:58220 ; RRID:Addgene_58220) |
| 参考文献部分: | Imaging intraorganellar Ca(2+) at subcellular resolution using CEPIA. Suzuki J, Kanemaru K, Ishii K, Ohkura M, Okubo Y, Iino M. Nat Commun. 2014 Jun 13;5:4153. doi: 10.1038/ncomms5153. 10.1038/ncomms5153 PubMed 24923787 |
买家导航
pCMVCEPIA4mt载体基本信息
pCMVCEPIA4mt载体图谱质粒图谱和多克隆位点信息
pCMVCEPIA4mt载体简介
pCMVCEPIA4mt载体序列
LOCUS pCMV CEPIA4mt 6404 bp ds-DNA circular SYN 11-MAR-2020
DEFINITION Green fluorescent indicator for calcium imaging in the mitochondria
(low affinity variant).
ACCESSION .
VERSION .
KEYWORDS pCMV CEPIA4mt
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6404)
AUTHORS Suzuki J, Kanemaru K, Ishii K, Ohkura M, Okubo Y, Iino M
TITLE Imaging intraorganellar Ca(2+) at subcellular resolution using
CEPIA.
JOURNAL Nat Commun. 2014 Jun 13;5:4153. doi: 10.1038/ncomms5153.
PUBMED 24923787
REFERENCE 2 (bases 1 to 6404)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Mar 11, 2020 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6404
/organism="synthetic DNA construct"
/mol_type="other DNA"
rep_origin complement(37..625)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(117..136)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(796..1656)
/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 1419..1438
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(1657..1761)
/gene="bla"
/label=AmpR promoter
enhancer 1826..2202
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 2204..2407
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 2358..2378
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
CDS 2660..2677
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 2681..2713
/codon_start=1
/product="leader peptide from bacteriophage T7 gene 10"
/label=T7 tag (gene 10 leader)
/note="promotes efficient translation in E. coli"
/translation="MASMTGGQQMG"
CDS 2717..2740
/codon_start=1
/product="Xpress(TM) epitope tag, including an enterokinase
recognition and cleavage site"
/label=Xpress(TM) tag
/translation="DLYDDDDK"
regulatory 2747..2756
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2768..2824
/codon_start=1
/product="calmodulin-binding peptide from avian smooth
muscle myosin light chain kinase"
/label=calmodulin-binding peptide
/translation="RRKWNKTGHAVRAIGRLSS"
primer_bind 3037..3058
/label=EGFP-C
/note="EGFP, forward primer"
primer_bind complement(3164..3185)
/label=EGFP-N
/note="EGFP, reverse primer"
CDS 3560..4000
/codon_start=1
/product="calcium-binding protein"
/label=calmodulin
/translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKDLGTVLRSLGQNPTEA
ELQDMINEVDADGNGTIDFPEFLTMMARKMKDTDSEEEIREAFRVWDKDGNGYISAAEL
RHVMTNLGEKLTDEEVDEMIREADIDGEGQVNYEEFVQMMTAK"
CDS 4010..4039
/codon_start=1
/product="Myc (human c-Myc proto-oncogene) epitope tag"
/label=Myc
/translation="EQKLISEEDL"
primer_bind complement(4075..4092)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 4081..4305
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 4351..4779
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4438..4457)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4648..4669
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(4788..4808)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 4793..5122
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 4973..5108
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 5035..5054
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 5148..5942
/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="MVEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(5202..5221)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 5812..5831
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 6116..6237
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(6153..6172)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 6207..6226
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(6270..6287)
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
61 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
121 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
181 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
241 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgta tgcacgaacc
301 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
361 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
421 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
481 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
541 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
601 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
661 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
721 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
781 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
841 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
901 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
961 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
1021 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
1081 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
1141 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
1201 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
1261 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
1321 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
1381 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
1441 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
1501 accgctgttg agatccagtt cgatgtaacc cactcgggca cccaactgat cttcagcatc
1561 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
1621 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
1681 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
1741 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcagatcgac
1801 ggatcgggag atcggtaccg aattcacatt gattattgag tagttattaa tagtaatcaa
1861 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
1921 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
1981 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
2041 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
2101 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
2161 ctacttggca gtacatctac ggttagtcat cgctattacc atagtgatgc ggttttggca
2221 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
2281 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
2341 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
2401 cagagctttc tggctaacta gagaacccac tgcttactgg cacgtgacca tgtccgtcct
2461 gacgccgctg ctgctgcggg gcttgacagg ctcggcccgg cggctcccag tgccgcgcgc
2521 caaaattcat tcactggggg accccatgag cgtgctcacc ccactcctgc tgcgggggct
2581 gaccggcagc gctaggcggc tgccagtccc gcgggccaag atccacagtc tcggcgatcc
2641 cggatccatg cggggttctc atcatcatca tcatcatggt atggctagca tgactggtgg
2701 acagcaaatg ggtcgggatc tgtacgacga tgacgataag gatctcgcca ccatggtcga
2761 ctcatcacgt cgtaagtgga ataagacagg tcacgcagtc agagctatag gtcggctgag
2821 ctcactcgag aacgtctata tcatggccga caagcagaag aacggcatca aggcgaactt
2881 caagatccgc cacaacatcg aggacggcgg cgtgcagctc gcctaccact accagcagaa
2941 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
3001 caaactttcg aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
3061 cgccgccggg atcactctcg gcatggacga gctgtacaag ggcggtaccg gagggagcat
3121 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
3181 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggt gagggcgatg ccacctacgg
3241 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
3301 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
3361 gcacgacttc ttcaagtccg ccatgcccga aggctacatc caggagcgca ccatcttctt
3421 caaggacgac ggctactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
3481 gaaccgcatc gtgctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
3541 gctggagtac aacacgcgtg accaactgac tgaagagcag atcgcagaat ttaaagaggc
3601 tttctcccta tttgacaagg acggggatgg gacaataaca accaaggatc tggggacggt
3661 gctgcggtct ctggggcaga accccacaga agcagagctc caggacatga tcaatgaagt
3721 agatgccgac ggtaatggca caatcgactt ccctgagttc ctgacaatga tggcaagaaa
3781 aatgaaagac acagacagtg aagaagaaat tcgcgaagcg ttccgtgtgt gggataagga
3841 tggcaatggc tacatcagtg cagcagagct tcgccacgtg atgacaaacc ttggagagaa
3901 gttaacagat gaagaggttg atgaaatgat cagggaagca gatatcgatg gagaaggtca
3961 ggtaaactac gaagagtttg tacaaatgat gacagcgaag gcggccgcag aacaaaaact
4021 catctcagaa gaggatctga atggggccgc atagtctaga agctcgctga tcagcctcga
4081 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
4141 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
4201 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
4261 gggaagacaa tagcaggcat gctggggatg gcccgggctc tatggcttct gaggcggaaa
4321 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
4381 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
4441 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
4501 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctctcc cccaaaaaac
4561 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
4621 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
4681 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
4741 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
4801 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
4861 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
4921 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
4981 cctaactccg cccagttccg cccattctcc gcccctaggc tgactaattt tttttattta
5041 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
5101 tggaggccta ggcttttgca aaaagctccc cccccgggag gtccacaatg gttgaacaag
5161 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
5221 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
5281 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactccag gacgaggcag
5341 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
5401 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
5461 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
5521 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
5581 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
5641 tcgcgccagc cgaactgttc gccaggctca aggcgcgtat gcccgacggc gaggatctcg
5701 tcgtgactca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
5761 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
5821 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
5881 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
5941 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
6001 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
6061 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
6121 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
6181 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
6241 tgtctgtata ccgtcgatct ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
6301 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
6361 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaa
//
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