| 质粒名称: | pET His6 MBP TEV LIC cloning vector (1M) |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 29656 |
| 存储实验室: | Scott Gradia |
|
载体骨架基本信息 |
|
| 载体名称: | pET |
| 载体出品公司: | -- |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | 6465 bp |
| 载体修饰: | -- |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | 低拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | -- |
| 物种来源: | -- |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | His6 MBP TEV（主干上的N终端） |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | LIC tag（克隆过程中销毁） |
| 3' 克隆位点: | LIC tag（克隆过程中销毁） |
| 5' 测序引物 | MBP forward (5'ggtcgtcagactgtcgatgaagcc) |
| 3' 测序引物 | T7 reverse |
|
文献引用方法 |
|
| 材料和方法部分: | pET His6 MBP TEV LIC cloning vector (1M) was a gift from Scott Gradia (Addgene plasmid # 29656 ; http://n2t.net/addgene:29656 ; RRID:Addgene_29656) |
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
质粒图谱.png)
载体简介
载体序列
LOCUS pET His6 MBP TEV LIC cloning vector (1M) 6462 bp ds-DNA circular SYN 12-MAR-2019
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pET His6 MBP TEV LIC cloning vector (1M)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6462)
TITLE LIC N-terminal fusion vectors for E. coli expression
REFERENCE 2 (bases 1 to 6462)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Mar 12, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6462
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 106..125
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind 209..228
/label=T7
/note="T7 promoter, forward primer"
promoter 209..227
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 228..252
/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 267..289
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 308..325
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 335..1435
/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
ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
WYAVRTAVINAASGRQTVDEALKDAQT"
primer_bind 1409..1432
/label=MBP-F
/note="Maltose binding protein, forward primer"
CDS 1448..1468
/codon_start=1
/product="tobacco etch virus (TEV) protease recognition and
cleavage site"
/label=TEV site
/translation="ENLYFQS"
CDS 1534..1551
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(1604..1622)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 1618..1665
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 1702..2157
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1789..1808)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 1999..2020
/label=F1ori-F
/note="F1 origin, forward primer"
CDS complement(2250..3065)
/codon_start=1
/gene="aph(3')-Ia"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin in bacteria or G418
(Geneticin(R)) in eukaryotes"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
primer_bind 2973..2992
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
primer_bind 3057..3076
/label=pENTR-R
/note="pENTR vectors, reverse primer"
rep_origin 3187..3775
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3676..3695
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 3929..3946
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 3961..4100
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(4026..4045)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 4186..4208
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(4202..4393)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(5202..6284)
/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 6246..6265
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(6285..6362)
/gene="lacI"
/label=lacI promoter
ORIGIN
1 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
61 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
121 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
181 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
241 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat ataccatggg
301 ttcttctcac catcaccatc accatggttc ttctatgaaa atcgaagaag gtaaactggt
361 aatctggatt aacggcgata aaggctataa cggtctcgct gaagtcggta agaaattcga
421 gaaagatacc ggaattaaag tcaccgttga gcatccggat aaactggaag agaaattccc
481 acaggttgcg gcaactggcg atggccctga cattatcttc tgggcacacg accgctttgg
541 tggctacgct caatctggcc tgttggctga aatcaccccg gacaaagcgt tccaggacaa
601 gctgtatccg tttacctggg atgccgtacg ttacaacggc aagctgattg cttacccgat
661 cgctgttgaa gcgttatcgc tgatttataa caaagatctg ctgccgaacc cgccaaaaac
721 ctgggaagag atcccggcgc tggataaaga actgaaagcg aaaggtaaga gcgcgctgat
781 gttcaacctg caagaaccgt acttcacctg gccgctgatt gctgctgacg ggggttatgc
841 gttcaagtat gaaaacggca agtacgacat taaagacgtg ggcgtggata acgctggcgc
901 gaaagcgggt ctgaccttcc tggttgacct gattaaaaac aaacacatga atgcagacac
961 cgattactcc atcgcagaag ctgcctttaa taaaggcgaa acagcgatga ccatcaacgg
1021 cccgtgggca tggtccaaca tcgacaccag caaagtgaat tatggtgtaa cggtactgcc
1081 gaccttcaag ggtcaaccat ccaaaccgtt cgttggcgtg ctgagcgcag gtattaacgc
1141 cgccagtccg aacaaagagc tggcaaaaga gttcctcgaa aactatctgc tgactgatga
1201 aggtctggaa gcggttaata aagacaaacc gctgggtgcc gtagcgctga agtcttacga
1261 ggaagagttg gcgaaagatc cacgtattgc cgccactatg gaaaacgccc agaaaggtga
1321 aatcatgccg aacatcccgc agatgtccgc tttctggtat gccgtgcgta ctgcggtgat
1381 caacgccgcc agcggtcgtc agactgtcga tgaagccctg aaagacgcgc agactaatgg
1441 gatcgaggaa aacctgtact tccaatccaa tattggaagt ggataacgga tccgaattcg
1501 agcgccgtcg acaagcttgc ggccgcactc gagcaccacc accaccacca ctgagatccg
1561 gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta
1621 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa aggaggaact
1681 atatccggat tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
1741 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
1801 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
1861 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
1921 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
1981 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
2041 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
2101 agctgattta acaaaaattt aacgcgaatt ttaacaaact agtaacgttt acaatttcag
2161 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
2221 caaatatgta tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac
2281 tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat
2341 gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg
2401 attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta
2461 tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc
2521 atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca
2581 tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg
2641 ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca
2701 tcaacaatgt tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg
2761 gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc
2821 ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg
2881 gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat
2941 cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa
3001 tcagcatcca tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg
3061 ctcataacac cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa
3121 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
3181 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
3241 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
3301 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
3361 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
3421 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
3481 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
3541 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
3601 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
3661 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
3721 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
3781 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
3841 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
3901 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
3961 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca atggtgcact
4021 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac
4081 gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
4141 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
4201 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag
4261 cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt
4321 tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt
4381 cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac
4441 cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac
4501 tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca
4561 ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc
4621 agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca
4681 gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt
4741 tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa
4801 ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg
4861 gggccgccat gccggcgata atggcctgct tctcgccgaa acgtttggtg gcgggaccag
4921 tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca
4981 tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct gccggcacct
5041 gtcctacgag ttgcatgata aagaagacag tcataagtgc ggcgacgata gtcatgcccc
5101 gcgcccaccg gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg
5161 gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt
5221 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
5281 tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg
5341 cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc
5401 aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta
5461 tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg
5521 cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgataccc
5581 tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt
5641 tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga
5701 cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg
5761 accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg
5821 ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca
5881 gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg
5941 agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc
6001 accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc
6061 gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt
6121 tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc
6181 cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag
6241 acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat
6301 tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg
6361 tccgggatct cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag
6421 gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca tg
//
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