| 质粒名称: | Flag-Rab5A |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 28043 |
| 存储实验室: | Qing Zhong |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA5/FRT/TO |
| 载体出品公司: | Invitrogen |
| 空载体大小: | 5137 bp |
| 完整质粒大小: | 5810bp |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | Hygromycin |
| 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: | Rab5A |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 647 bp |
| 标签/融合蛋白: | 3xFlag (C terminal on insert) |
| 克隆方法: | 限制性内切酶 |
| 5’测序引物 | CMV-F |
| 3’测序引物 | BGH-rev |
|
文献引用方法 |
|
| 材料和方法部分: | Flag-Rab5A was a gift from Qing Zhong (Addgene plasmid # 28043 ; http://n2t.net/addgene:28043 ; RRID:Addgene_28043) |
| 参考文献部分: | Rubicon controls endosome maturation as a Rab7 effector. Sun Q, Westphal W, Wong KN, Tan I, Zhong Q. Proc Natl Acad Sci U S A. 2010 Nov 9. 107(45):19338-43. 10.1073/pnas.1010554107 PubMed 20974968 |
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 Flag-Rab5A 5810 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS Flag-Rab5A
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5810)
AUTHORS Sun Q, Westphal W, Wong KN, Tan I, Zhong Q
TITLE Rubicon controls endosome maturation as a Rab7 effector.
JOURNAL Proc Natl Acad Sci U S A. 2010 Nov 9. 107(45):19338-43.
PUBMED 20974968
REFERENCE 2 (bases 1 to 5810)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofengcom
FEATURES Location/Qualifiers
source 1..5810
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 92..471
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 472..675
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 626..646
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
protein_bind 677..695
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 698..716
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
primer_bind 726..750
/label=LNCX
/note="Human CMV promoter, forward primer"
CDS 1513..1578
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
primer_bind complement(1619..1636)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 1625..1849
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind complement(1982..2001)
/label=F1ori-R
/note="F1 origin, reverse primer"
protein_bind 2133..2180
/label=FRT
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2u plasmid"
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
CDS 2188..3210
/codon_start=1
/gene="aph(4)-Ia"
/product="aminoglycoside phosphotransferase from E. coli"
/label=HygR
/note="confers resistance to hygromycin"
/translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
"
polyA_signal 3340..3461
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(3377..3396)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 3431..3450
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(3510..3526)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(3510..3526)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(3523..3545)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 3534..3550
/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(3558..3588)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 3603..3624
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(3741..3758)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3912..4500)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3992..4011)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(4671..5531)
/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 5294..5313
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(5532..5636)
/gene="bla"
/label=AmpR promoter
primer_bind complement(5711..5730)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 aggcaaggct tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg
61 cttcgcgatg tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag
121 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
181 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
241 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
301 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
361 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
421 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
481 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
541 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
601 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
661 tatataagca gagctctccc tatcagtgat agagatctcc ctatcagtga tagagatcgt
721 cgacgagctc gtttagtgaa ccgtcagatc gcctggagac gccatccacg ctgttttgac
781 ctccatagaa gacaccggga ccgatccagc ctccggactc tagcgtttaa acttaagctt
841 ggtaccgagc tcggatccat ggctagtcga ggcgcaacaa gacccaacgg gccaaatact
901 ggaaataaaa tatgccagtt caaactagta cttctgggag agtccgctgt tggcaaatca
961 agcctagtgc ttcgttttgt gaaaggccaa tttcatgaat ttcaagagag taccattggg
1021 gctgcttttc taacccaaac tgtatgtctt gatgacacta cagtaaagtt tgaaatatgg
1081 gatacagctg gtcaagaacg ataccatagc ctagcaccaa tgtactacag aggagcacaa
1141 gcagccatag ttgtatatga tatcacaaat gaggagtcct ttgcaagagc aaaaaattgg
1201 gttaaagaac ttcagaggca agcaagtcct aacattgtaa tagctttatc gggaaacaag
1261 gccgacctag caaataaaag agcagtagat ttccaggaag cacagtccta tgcagatgac
1321 aatagtttat tattcatgga gacatccgct aaaacatcaa tgaatgtaaa tgaaatattc
1381 atggcaatag ctaaaaaatt gccaaagaat gaaccacaaa atccaggagc aaattctgcc
1441 agaggaagag gagtagacct taccgaaccc acacaaccaa ccaggaatca gtgttgtagt
1501 aacggcggcc gcgactacaa agaccatgac ggtgattata aagatcatga catcgactac
1561 aaggatgacg atgacaagtg actcgagtct agagggcccg tttaaacccg ctgatcagcc
1621 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
1681 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
1741 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
1801 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
1861 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
1921 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
1981 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
2041 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
2101 aaacttgatt agggtgatgg ttcacgtacc tagaagttcc tattccgaag ttcctattct
2161 ctagaaagta taggaacttc cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag
2221 aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa
2281 gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc
2341 tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc
2401 ccgattccgg aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc
2461 cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg
2521 cagccggtcg cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg
2581 ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc
2641 gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg
2701 tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg
2761 cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca
2821 gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc
2881 ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg
2941 catccggagc ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac
3001 caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga
3061 tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga
3121 agcgcggccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc
3181 cccagcactc gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc
3241 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
3301 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
3361 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
3421 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
3481 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
3541 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
3601 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
3661 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
3721 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
3781 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
3841 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
3901 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
3961 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
4021 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
4081 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
4141 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
4201 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
4261 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
4321 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
4381 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
4441 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
4501 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
4561 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
4621 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
4681 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
4741 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
4801 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
4861 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
4921 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
4981 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
5041 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
5101 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
5161 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
5221 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
5281 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
5341 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
5401 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
5461 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
5521 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
5581 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
5641 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
5701 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
5761 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca
//
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