| 质粒名称: | Flag-miniTurbo |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 124647 |
| 存储实验室: | FENG-QIAN Li, Ken-Ichi Takemaru |
|
载体骨架基本信息 |
|
| 载体名称: | pCD-betaG-FLAG |
| 载体类型: | 哺乳细胞表达 |
| 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: | miniTurbo |
| 启动子: | CMV |
| 标签/融合蛋白: | Flag (N terminal on backbone) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | BglII (destroyed during cloning) |
| 3’克隆位点: | BglII (未破坏) |
| 5’测序引物 | GCGTGCCTAATGGGAGGTCT |
|
文献引用方法 |
|
| 材料和方法部分: | Flag-miniTurbo was a gift from FENG-QIAN Li & Ken-Ichi Takemaru (Addgene plasmid # 124647 ; http://n2t.net/addgene:124647 ; RRID:Addgene_124647) |
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-miniTurbo 4910 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Expresses Flag-miniTurbo fusion for proximity labeling.
ACCESSION .
VERSION .
KEYWORDS Flag-miniTurbo
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4910)
TITLE Expresses Flag-miniTurbo fusion for proximity labeling
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 4910)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..4910
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..978
/label=CMV IE94 promoter
/note="enhancer/promoter region of simian cytomegalovirus
major immediate early transcription unit IE94"
promoter 1013..1031
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 1013..1030
/label=SP6
/note="SP6 promoter, forward primer"
CDS 1069..1092
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
CDS 1099..1869
/codon_start=1
/product="N-terminally truncated mutant of the E. coli
biotin ligase BioID engineered for improved catalytic
efficiency (Branon et al., 2018)"
/label=miniTurbo
/translation="IPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSGDACI
AEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKLGADK
VRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGWITLQ
EAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIGDKEIF
GISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
promoter complement(1918..1935)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
polyA_signal 1940..2074
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1964..1983)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 2018..2037
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind complement(2187..2207)
/label=T3
/note="T3 promoter, forward primer"
promoter complement(2187..2205)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(2226..2242)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(2226..2242)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(2239..2261)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 2250..2266
/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(2274..2304)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2319..2340
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(2457..2474)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2628..3216)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2708..2727)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(3387..4247)
/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 4010..4029
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(4248..4352)
/gene="bla"
/label=AmpR promoter
rep_origin complement(4378..4833)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4515..4536)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 4727..4746
/label=F1ori-R
/note="F1 origin, reverse primer"
ORIGIN
1 gccaattcaa tatggcgtat atggactcat gccaattcaa tatggtggat ctggacctgt
61 gccaattcaa tatggcgtat atggactcgt gccaattcaa tatggtggat ctggacccca
121 gccaattcaa tatggcggac ttggcaccat gccaattcaa tatggcggac ctggcactgt
181 gccaactggg gaggggtcta cttggcacgg tgccaagttt gaggaggggt cttggccctg
241 tgccaagtcc gccatattga attggcatgg tgccaataat ggcggccata ttggctatat
301 gccaggatca atatataggc aatatccaat atggccctat gccaatatgg ctattggcca
361 ggttcaatac tatgtattgg ccctatgcca tatagtattc catatatggg ttttcctatt
421 gacgtagata gcccctccca atgggcggtc ccatatacca tatatggggc ttcctaatac
481 cgcccatagc cactccccca ttgacgtcaa tggtctctat atatggtctt tcctattgac
541 gtcatatggg cggtcctatt gacgtatatg gcgcctcccc cattgacgtc aattacggta
601 aatggcccgc ctggctcaat gcccattgac gtcaatagga ccacccacca ttgacgtcaa
661 tgggatggct cattgcccat tcatatccgt tctcacgccc cctattgacg tcaatgacgg
721 taaatggccc acttggcagt acatcaatat ctattaatag taacttggca agtacattac
781 tattggaagt acgccagggt acattggcag tactcccatt gacgtcaatg gcggtaaatg
841 gcccgcgatg gctgccaagt acatccccat tgacgtcaat ggggaggggc aatgacgcaa
901 atgggcgttc cattgacgta aatgggcggt aggcgtgcct aatgggaggt ctatataagc
961 aatgctcgtt tagggaaccg ccattctgcc tggggacgtc ggagcaagct tgatttaggt
1021 gacactatag aatacaagct acttgttctt tttgcaggat ccaccatgga ttacaaggat
1081 gacgacgata agatccgcat cccgctgctg aacgctaaac agattctggg acagctggac
1141 ggcgggagcg tggcagtcct gcctgtggtc gactccacca atcagtacct gctggatcga
1201 atcggcgagc tgaagagtgg ggatgcttgc attgcagaat atcagcaggc agggagagga
1261 agcagaggga ggaaatggtt ctctcctttt ggagctaacc tgtacctgag tatgttttgg
1321 cgcctgaagc ggggaccagc agcaatcggc ctgggcccgg tcatcggaat tgtcatggca
1381 gaagcgctgc gaaagctggg agcagacaag gtgcgagtca aatggcccaa tgacctgtat
1441 ctgcaggata gaaagctggc aggcatcctg gtggagctgg ccggaataac aggcgatgct
1501 gcacagatcg tcattggcgc cgggattaac gtggctatga ggcgcgtgga ggaaagcgtg
1561 gtcaatcagg gctggatcac actgcaggaa gcagggatta acctggacag gaatactctg
1621 gccgctatgc tgatccgaga gctgcgggca gccctggaac tgttcgagca ggaaggcctg
1681 gctccatatc tgtcacggtg ggagaagctg gataacttca tcaatagacc cgtgaagctg
1741 atcattgggg acaaagagat tttcgggatt agccggggga ttgataaaca gggagccctg
1801 ctgctggaac aggacggagt tatcaaaccc tggatgggcg gagaaatcag tctgcggtct
1861 gccgaaaagc tgcagggatc tcagatctcg aattcaaggc ctctcgagcc tctagaacta
1921 tagtgagtcg tattacgtag atccagacat gataagatac attgatgagt ttggacaaac
1981 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
2041 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
2101 gtttcaggtt cagggggagg tgtgggaggt tttttaattc gcggccgcgg cgccaatgca
2161 ttgggcccgg tacccagctt ttgttccctt tagtgagggt taattgcgcg cttggcgtaa
2221 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
2281 cgagccggga gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
2341 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
2401 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
2461 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
2521 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
2581 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
2641 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
2701 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
2761 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
2821 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
2881 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
2941 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
3001 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
3061 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
3121 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
3181 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
3241 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
3301 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
3361 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
3421 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
3481 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
3541 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
3601 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
3661 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
3721 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
3781 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
3841 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
3901 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
3961 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
4021 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
4081 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
4141 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
4201 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
4261 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
4321 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa
4381 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
4441 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
4501 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
4561 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
4621 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
4681 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
4741 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
4801 ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg ctgcgcaact
4861 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagtcgacc
//
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