| 质粒名称: | pMal-T-Avi-His/BirA |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 102962 |
| 存储实验室: | Tonia Rex |
|
载体骨架基本信息 |
|
| 载体名称: | pMAL-c5X |
| 载体出品公司: | NEB |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 5675 bp |
| 载体类型: | 细菌表达 |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 30 ℃ |
|
插入基因信息 |
|
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | Kpn 1(未破坏) |
| 3' 克隆位点: | Nhe 1 (未破坏) |
| 5' 测序引物 | GGTCGTCAGACTGTCGATGAAGCC |
| 3' 测序引物 | TGTCCTACTCAGGAGAGCGTTCAC |
|
文献引用方法 |
|
| 材料和方法部分: | pMal-T-Avi-His/BirA was a gift from Tonia Rex (Addgene plasmid # 102962 ; http://n2t.net/addgene:102962 ; RRID:Addgene_102962) |
| 参考文献: | Piezo1 and Piezo2 are essential components of distinct mechanically activated cation channels. Coste B, Mathur J, Schmidt M, Earley TJ, Ranade S, Petrus MJ, Dubin AE, Patapoutian A. Science. 2010 Oct 1;330(6000):55-60. doi: 10.1126/science.1193270. Epub 2010 Sep 2. 10.1126/science.1193270 PubMed 20813920 |
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 pMal-T-Avi-His/BirA 7292 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Vector for in vivo biotinylation of a fusion protein in bacteria..
ACCESSION .
VERSION .
KEYWORDS pMal-T-Avi-His/BirA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7292)
AUTHORS Tonia Rex
TITLE Tonia Rex lab plasmids
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 7292)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7292
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 85..107
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(101..292)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
promoter 788..865
/gene="lacI (mutant)"
/label=lacIq promoter
/note="In the lacIq allele, a single base change in the
promoter boosts expression of the lacI gene about 10-fold."
CDS 866..1948
/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 complement(885..904)
/label=LacI-R
/note="LacI, reverse primer"
promoter 2191..2219
/label=tac promoter
/note="strong E. coli promoter; hybrid between the trp and
lac UV5 promoters"
protein_bind 2227..2243
/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)."
primer_bind 2232..2254
/label=M13/pUC Reverse
/note="In lacZ gene"
CDS 2313..3413
/codon_start=1
/gene="malE (mutated)"
/product="maltose binding protein from E. coli"
/label=MBP
/note="This version of the gene does not encode a signal
sequence, so MBP will remain in the cytosol."
/translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
ENYLLTDEGLEAVNKDKPLGAVALKSYEEELVKDPRIAATMENAQKGEIMPNIPQMSAF
WYAVRTAVINAASGRQTVDEALKDAQT"
primer_bind 3387..3410
/label=MBP-F
/note="Maltose binding protein, forward primer"
CDS 3462..3473
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
CDS 3551..3571
/codon_start=1
/product="tobacco etch virus (TEV) protease recognition and
cleavage site"
/label=TEV site
/translation="EDLYFQS"
CDS 3638..3655
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 3664..3687
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
terminator 3699..3785
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 3877..3904
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"
promoter 3932..4022
/gene="bla"
/label=AmpR promoter
CDS 4023..4883
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(4241..4260)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 4971..5559
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 5460..5479
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 5725..5744
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
protein_bind 5847..5871
/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 5886..5908
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 5915..6877
/codon_start=1
/gene="birA"
/product="E. coli biotin protein ligase"
/label=BirA
/translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
CDS 6916..6933
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(6986..7004)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 7000..7047
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
primer_bind 7177..7194
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
61 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
121 caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtgc agcgattcac
181 agatgtctgc ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct
241 ggcttctgat aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct
301 ccgtgtaagg gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc
361 tcacgatacg ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac
421 aactggcggt atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct
481 tcgttaatac agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc
541 ggaacataat ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac
601 cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg
661 ttcgctcgcg tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc
721 gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg ccaggaccca acgctgcccg
781 aaattccgac accatcgaat ggtgcaaaac ctttcgcggt atggcatgat agcgcccgga
841 agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta
901 tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc
961 gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt
1021 ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc
1081 cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc
1141 cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca
1201 caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga
1261 tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga
1321 ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga
1381 gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt
1441 ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc
1501 gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat
1561 gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg
1621 cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatattt cggtagtggg
1681 atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga
1741 ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc
1801 ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc
1861 caatacacaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
1921 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc
1981 attaggcaca attctcatgt ttgacagctt atcatcgact gcacggtgca ccaatgcttc
2041 tggcgtcagg cagccatcgg aagctgtggt atggctgtgc aggtcgtaaa tcactgcata
2101 attcgtgtcg ctcaaggcgc actcccgttc tggataatgt tttttgcgcc gacatcataa
2161 cggttctggc aaatattctg aaatgagctg ttgacaatta atcatcggct cgtataatgt
2221 gtggaattgt gagcggataa caatttcaca caggaaacag ccagtccgtt taggtgtttt
2281 cacgagcaat tgaccaacaa ggaccataga ttatgaaaat cgaagaaggt aaactggtaa
2341 tctggattaa cggcgataaa ggctataacg gtctcgctga agtcggtaag aaattcgaga
2401 aagataccgg aattaaagtc accgttgagc atccggataa actggaagag aaattcccac
2461 aggttgcggc aactggcgat ggccctgaca ttatcttctg ggcacacgac cgctttggtg
2521 gctacgctca atctggcctg ttggctgaaa tcaccccgga caaagcgttc caggacaagc
2581 tgtatccgtt tacctgggat gccgtacgtt acaacggcaa gctgattgct tacccgatcg
2641 ctgttgaagc gttatcgctg atttataaca aagatctgct gccgaacccg ccaaaaacct
2701 gggaagagat cccggcgctg gataaagaac tgaaagcgaa aggtaagagc gcgctgatgt
2761 tcaacctgca agaaccgtac ttcacctggc cgctgattgc tgctgacggg ggttatgcgt
2821 tcaagtatga aaacggcaag tacgacatta aagacgtggg cgtggataac gctggcgcga
2881 aagcgggtct gaccttcctg gttgacctga ttaaaaacaa acacatgaat gcagacaccg
2941 attactccat cgcagaagct gcctttaata aaggcgaaac agcgatgacc atcaacggcc
3001 cgtgggcatg gtccaacatc gacaccagca aagtgaatta tggtgtaacg gtactgccga
3061 ccttcaaggg tcaaccatcc aaaccgttcg ttggcgtgct gagcgcaggt attaatgccg
3121 ccagtccgaa caaagagctg gcaaaagagt tcctcgaaaa ctatctgctg actgatgaag
3181 gtctggaagc ggttaataaa gacaaaccgc tgggtgccgt agcgctgaag tcttacgagg
3241 aagagttggt gaaagatccg cgtattgccg ccactatgga aaacgcccag aaaggtgaaa
3301 tcatgccgaa catcccgcag atgtccgctt tctggtatgc cgtgcgtact gcggtgatca
3361 acgccgccag cggtcgtcag actgtcgatg aagccctgaa agacgcgcag actaattcga
3421 gttctaacaa caacaacaat aacaataaca acaacctcgg gatcgaggga aggatttcac
3481 atatgtccat gggcggccgc gatatcgtcg acaagaggtg atatcgatcg gagctctcga
3541 gccaaccact gaggatctgt actttcagag cgataacgat ggatccctag gactgaacga
3601 catcttcgaa gctcagaaaa tcgaatggca cggatcccat catcaccatc accactagaa
3661 ttcgattaca aggatgacga cgataagtaa ataagcttca aataaaacga aaggctcagt
3721 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga
3781 caaatccgcc gggagcggat ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag
3841 gacgcccgcc ataaactgcc aggcatcaaa ttaagcagaa ggccatcctg acggatggcc
3901 tttttgcgtt tctacaaact ctttcggtcc gttgtttatt tttctaaata cattcaaata
3961 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
4021 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
4081 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
4141 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
4201 ccgaagaacg tttcccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
4261 cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
4321 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
4381 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
4441 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
4501 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
4561 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
4621 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
4681 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
4741 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
4801 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
4861 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
4921 atttccttag gactgagcgt caaccccgta gaaaagatca aaggatcttc ttgagatcct
4981 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
5041 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
5101 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
5161 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
5221 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
5281 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
5341 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
5401 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
5461 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
5521 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
5581 ttacggttcc tggccttttg ctggcctttt gctcacatgt tccggtaccg cgcccaacag
5641 tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa
5701 gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc
5761 tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga tctcgatccc
5821 gcgaaattaa tacgactcac tagaggggaa ttgtgagcgg ataacaattc ccctctagaa
5881 ataattttgt ttaactttaa gaaggagata taccatgaag gataacaccg tgccactgaa
5941 attgattgcc ctgttagcga acggtgaatt tcactctggc gagcagttgg gtgaaacgct
6001 gggaatgagc cgggcggcta ttaataaaca cattcagaca ctgcgtgact ggggcgttga
6061 tgtctttacc gttccgggta aaggatacag cctgcctgag cctatccagt tacttaatgc
6121 taaacagata ttgggtcagc tggatggcgg tagtgtagcc gtgctgccag tgattgactc
6181 cacgaatcag taccttcttg atcgtatcgg agagcttaaa tcgggcgatg cttgcattgc
6241 agaataccag caggctggcc gtggtcgccg gggtcggaaa tggttttcgc cttttggcgc
6301 aaacttatat ttgtcgatgt tctggcgtct ggaacaaggc ccggcggcgg cgattggttt
6361 aagtctggtt atcggtatcg tgatggcgga agtattacgc aagctgggtg cagataaagt
6421 tcgtgttaaa tggcctaatg acctctatct gcaggatcgc aagctggcag gcattctggt
6481 ggagctgact ggcaaaactg gcgatgcggc gcaaatagtc attggagccg ggatcaacat
6541 ggcaatgcgc cgtgttgaag agagtgtcgt taatcagggg tggatcacgc tgcaggaagc
6601 ggggatcaat ctcgatcgta atacgttggc ggccatgcta atacgtgaat tacgtgctgc
6661 gttggaactc ttcgaacaag aaggattggc accttatctg tcgcgctggg aaaagctgga
6721 taattttatt aatcgcccag tgaaacttat cattggtgat aaagaaatat ttggcatttc
6781 acgcggaata gacaaacagg gggctttatt acttgagcag gatggaataa taaaaccctg
6841 gatgggcggt gaaatatccc tgcgtagtgc agaaaaaggt aagctttgat gaggactagt
6901 gcggccgcac tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga
6961 aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc
7021 tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg atgctagcac
7081 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
7141 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
7201 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
7261 taaggtgcac tgtgactggg tcatggctgc gc
//
