| 质粒名称: | pDEST/LIfeAct-mCherry-N1 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 40908 |
| 存储实验室: | Robin Shaw |
|
载体骨架基本信息 |
|
| 载体名称: | pmcherry-N1 |
| 载体出品公司: | Clontech |
| 空载体大小: | 4722 bp |
| 完整质粒大小: | 4831 bp |
| 载体修饰: | Gateway reading frame cassette c.1 inserted between Afe I and Sma I sites of MCS. |
| 载体类型: | 哺乳动物表达 |
| 筛选标记: | 新霉素（G418筛选） |
| 载体抗性: | 卡那霉素，50μg/mL |
| 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: | LifeAct |
| 物种来源: | 酿酒酵母（芽苗酵母） |
| 插入大小: | 51 bp |
| 基因突变: | 无终止密码子产生c端融合 |
| GenBank编号 | -- |
| 启动子: | CMV |
| 标签/融合蛋白: | mCherry（主干上的C终端） |
| 克隆方法: | Gateway Cloning |
| 5’克隆位点: | -- |
| 3’克隆位点: | -- |
| 5’测序引物 | CMV fwd |
| 3’测序引物 | mCherry修订版 |
|
文献引用方法 |
|
| 材料和方法部分: | pDEST/LIfeAct-mCherry-N1 was a gift from Robin Shaw (Addgene plasmid # 40908 ; http://n2t.net/addgene:40908 ; RRID:Addgene_40908) |
| 参考文献部分: | Actin cytoskeleton rest stops regulate anterograde traffic of connexin 43 vesicles to the plasma membrane. Smyth JW, Vogan JM, Buch PJ, Zhang SS, Fong TS, Hong TT, Shaw RM. Circ Res. 2012 Mar 30;110(7):978-89. Epub 2012 Feb 9. 10.1161/CIRCRESAHA.111.257964 PubMed 22328533 |
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 pDEST/LIfeAct-mCherry-N1 4831 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pDEST/LIfeAct-mCherry-N1
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4831)
AUTHORS Smyth JW, Vogan JM, Buch PJ, Zhang SS, Fong TS, Hong TT, Shaw RM
TITLE Actin cytoskeleton rest stops regulate anterograde traffic of
connexin 43 vesicles to the plasma membrane.
JOURNAL Circ Res. 2012 Mar 30;110(7):978-89. Epub 2012 Feb 9.
PUBMED 22328533
REFERENCE 2 (bases 1 to 4831)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..4831
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 54..357
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 358..561
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 512..532
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
protein_bind 595..619
/gene="mutant version of attB"
/label=attB1
/bound_moiety="BP Clonase(TM)"
/note="recombination site for the Gateway(R) BP reaction"
CDS 649..699
/codon_start=1
/product="peptide that binds filamentous actin (Riedl et
al., 2008)"
/label=Lifeact
/note="first 17 amino acids of budding yeast Abp140"
/translation="MGVADLIKKFESISKEE"
protein_bind complement(713..737)
/gene="mutant version of attB"
/label=attB2
/bound_moiety="BP Clonase(TM)"
/note="recombination site for the Gateway(R) BP reaction"
regulatory 751..760
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 757..1467
/codon_start=1
/product="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
/label=mCherry
/note="mammalian codon-optimized"
/translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
EGRHSTGGMDELYK"
primer_bind complement(902..920)
/label=mCherry-R
/note="mCherry, reverse primer"
primer_bind 1170..1189
/label=mCherry-F
/note="mCherry, forward primer"
polyA_signal 1612..1733
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1649..1668)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 1703..1722
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
rep_origin complement(1740..2195)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1877..1898)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 2089..2108
/label=F1ori-R
/note="F1 origin, reverse primer"
promoter 2222..2326
/gene="bla"
/label=AmpR promoter
promoter 2328..2685
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
primer_bind complement(2351..2371)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 2536..2671
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 2598..2617
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 2720..3514
/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(2774..2793)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 3384..3403
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
primer_bind complement(3702..3721)
/label=TK-pA-R
/note="Thymidine kinase polyA, reverse primer"
polyA_signal 3746..3793
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
rep_origin 4122..4710
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 4611..4630
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
61 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
121 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
181 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
241 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
301 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg
361 atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca
421 agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt
481 ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg
541 gaggtctata taagcagagc tggtttagtg aaccgtcaga tccgctagca tcaaacaagt
601 ttgtacaaaa aagcaggctt taaaggaacc aattcagtcg actggatcat gggcgtggcc
661 gacctgatca agaagttcga gagcatcagc aaggaggagt cgagatatct agacccagct
721 ttcttgtaca aagtggttcg atgggatccc gccaccatgg tgagcaaggg cgaggaggat
781 aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac
841 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc
901 gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat cctgtcccct
961 cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg
1021 aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
1081 gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag
1141 ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac catgggctgg
1201 gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga gatcaagcag
1261 aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac ctacaaggcc
1321 aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga catcacctcc
1381 cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg ccactccacc
1441 ggcggcatgg acgagctgta caagtaagaa ttcgaagctg tacaagtaaa gcggccgcga
1501 ctctagatca taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc
1561 ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt
1621 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
1681 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta
1741 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
1801 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
1861 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
1921 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
1981 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
2041 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
2101 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
2161 acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt
2221 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
2281 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag
2341 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
2401 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
2461 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
2521 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
2581 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
2641 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac
2701 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
2761 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
2821 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
2881 cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg
2941 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
3001 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
3061 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
3121 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
3181 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
3241 caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
3301 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
3361 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
3421 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
3481 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
3541 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
3601 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
3661 ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca
3721 ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt
3781 gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca
3841 ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca
3901 agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc
3961 tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
4021 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
4081 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
4141 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
4201 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
4261 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
4321 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
4381 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
4441 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
4501 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
4561 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
4621 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
4681 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
4741 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
4801 gataaccgta ttaccgccat gcattagtta t
//
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