| 质粒名称: | miniTurbo-His6_pET21a |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 107178 |
| 存储实验室: | Alice Ting |
|
载体骨架基本信息 |
|
| 载体名称: | pET21a |
| 载体类型: | 细菌细胞表达 |
| 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 (BirA mutant) |
| 基因突变: | aa1-63 deleted; Q65P, I87V, R118S, E140K, Q141R, S150G, L151P V160A, T192A, K194I, M209V, I305V |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | 未知 |
| 3’克隆位点: | 未知 |
|
文献引用方法 |
|
| 材料和方法部分: | miniTurbo-His6_pET21a was a gift from Alice Ting (Addgene plasmid # 107178 ; http://n2t.net/addgene:107178 ; RRID:Addgene_107178) |
| 参考文献部分: | Efficient proximity labeling in living cells and organisms with TurboID. Branon TC, Bosch JA, Sanchez AD, Udeshi ND, Svinkina T, Carr SA, Feldman JL, Perrimon N, Ting AY. Nat Biotechnol. 2018 Aug 20. pii: nbt.4201. doi: 10.1038/nbt.4201. 10.1038/nbt.4201 PubMed 30125270 |
买家导航
miniTurbo-His6_pET21a载体质粒基本信息
miniTurbo-His6_pET21a质粒图谱载体图谱和miniTurbo-His6_pET21a载体序列质粒序列多克隆位点信息
miniTurbo-His6_pET21a质粒载体简介
miniTurbo-His6_pET21a质粒序列载体序列
LOCUS miniTurbo-His6_pET21a 6200 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION expresses His6-tagged miniTurbo in bacteria.
ACCESSION .
VERSION .
KEYWORDS miniTurbo-His6_pET21a
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6200)
AUTHORS Branon TC, Bosch JA, Sanchez AD, Udeshi ND, Svinkina T, Carr SA,
Feldman JL, Perrimon N, Ting AY
TITLE Efficient proximity labeling in living cells and organisms with
TurboID.
JOURNAL Nat Biotechnol. 2018 Aug 20. pii: nbt.4201. doi: 10.1038/nbt.4201.
PUBMED 30125270
REFERENCE 2 (bases 1 to 6200)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..6200
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 48..67
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind 151..170
/label=T7
/note="T7 promoter, forward primer"
promoter 151..169
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 170..194
/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)."
RBS 209..231
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 239..271
/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 290..1060
/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"
CDS 1082..1099
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(1152..1170)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 1166..1213
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 1250..1705
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1337..1356)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 1547..1568
/label=F1ori-F
/note="F1 origin, forward primer"
promoter 1731..1835
/gene="bla"
/label=AmpR promoter
CDS 1836..2696
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(2054..2073)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 2867..3455
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3356..3375
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 3609..3626
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 3641..3780
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(3706..3725)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 3866..3888
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(3882..4073)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(4882..5964)
/codon_start=1
/gene="lacI"
/product="lac repressor"
/label=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)."
/translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
primer_bind 5926..5945
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(5965..6042)
/gene="lacI"
/label=lacI promoter
ORIGIN
1 aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg
61 atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
121 tagaggatcg agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag
181 cggataacaa ttcccctcta gaaataattt tgtttaactt taagaaggag atatacatat
241 ggctagcatg actggtggac agcaaatggg tcgcggatcc gaattcatga tcccgctgct
301 gaacgctaaa cagattctgg gacagctgga cggcgggagc gtggcagtcc tgcctgtggt
361 cgactccacc aatcagtacc tgctggatcg aatcggcgag ctgaagagtg gggatgcttg
421 cattgcagaa tatcagcagg cagggagagg aagcagaggg aggaaatggt tctctccttt
481 tggagctaac ctgtacctga gtatgttttg gcgcctgaag cggggaccag cagcaatcgg
541 cctgggcccg gtcatcggaa ttgtcatggc agaagcgctg cgaaagctgg gagcagacaa
601 ggtgcgagtc aaatggccca atgacctgta tctgcaggat agaaagctgg caggcatcct
661 ggtggagctg gccggaataa caggcgatgc tgcacagatc gtcattggcg ccgggattaa
721 cgtggctatg aggcgcgtgg aggaaagcgt ggtcaatcag ggctggatca cactgcagga
781 agcagggatt aacctggaca ggaatactct ggccgctatg ctgatccgag agctgcgggc
841 agccctggaa ctgttcgagc aggaaggcct ggctccatat ctgtcacggt gggagaagct
901 ggataacttc atcaatagac ccgtgaagct gatcattggg gacaaagaga ttttcgggat
961 tagccggggg attgataaac agggagccct gctgctggaa caggacggag ttatcaaacc
1021 ctggatgggc ggagaaatca gtctgcggtc tgccgaaaag aagcttgcgg ccgcactcga
1081 gcaccaccac caccaccact gagatccggc tgctaacaaa gcccgaaagg aagctgagtt
1141 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
1201 gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt
1261 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
1321 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
1381 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
1441 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
1501 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
1561 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
1621 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
1681 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
1741 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
1801 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
1861 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
1921 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
1981 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
2041 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
2101 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
2161 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
2221 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
2281 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
2341 aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc
2401 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
2461 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
2521 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
2581 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
2641 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
2701 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
2761 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
2821 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
2881 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
2941 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
3001 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
3061 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
3121 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
3181 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
3241 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
3301 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
3361 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
3421 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
3481 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
3541 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
3601 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc
3661 ttacgcatct gtgcggtatt tcacaccgca atggtgcact ctcagtacaa tctgctctga
3721 tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg
3781 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
3841 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
3901 tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca
3961 cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc
4021 tggcttctga taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc
4081 tccgtgtaag ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg
4141 ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa
4201 caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc
4261 ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc
4321 cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa
4381 ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac
4441 gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc
4501 cgggtcctca acgacaggag cacgatcatg cgcacccgtg gggccgccat gccggcgata
4561 atggcctgct tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg
4621 gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag
4681 cggtcctcgc cgaaaatgac ccagagcgct gccggcacct gtcctacgag ttgcatgata
4741 aagaagacag tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg
4801 actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg agtgagctaa
4861 cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
4921 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt
4981 ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg
5041 agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat
5101 ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga
5161 gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat
5221 ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt
5281 ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg
5341 attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa
5401 tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag
5461 tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc
5521 aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc
5581 cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc
5641 tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg
5701 atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga
5761 ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg
5821 aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg
5881 gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac
5941 atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta
6001 tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc
6061 ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg
6121 ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc
6181 ctgccaccat acccacgccg
//
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