| 质粒名称: | pET28-MHL |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 26096 |
| 存储实验室: | Cheryl Arrowsmith |
|
载体骨架基本信息 |
|
| 载体名称: | pET28-MHL |
| 载体出品公司: | Structural Genomics Consortium |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | 7325 bp |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | 高拷贝 |
| 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 | T7-lacO |
| 标签/融合蛋白: | 具有TEV裂解的六组氨酸标签（主链上的N端） |
| 克隆方法: | -- |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | T7 |
| 3' 测序引物 | T7-term |
|
文献引用方法 |
|
| 材料和方法部分: | pET28-MHL was a gift from Cheryl Arrowsmith (Addgene plasmid # 26096 ; http://n2t.net/addgene:26096 ; RRID:Addgene_26096) |
买家导航
pET28-MHL载体 pET28MHL质粒基本信息
pET28-MHL质粒图谱 pET28MHL载体图谱和pET28-MHL载体序列 pET28MHL质粒序列多克隆位点信息
pET28-MHL质粒 pET28MHL载体简介
pET28-MHL质粒序列 pET28MHL载体序列
LOCUS pET28-MHL 7325 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION SGC Empty backbone for bacterial expression.
ACCESSION .
VERSION .
KEYWORDS pET28-MHL
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7325)
TITLE Cloning Vectors
REFERENCE 2 (bases 1 to 7325)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7325
/organism="synthetic DNA construct"
/mol_type="other DNA"
rep_origin 12..467
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(99..118)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 309..330
/label=F1ori-F
/note="F1 origin, forward primer"
CDS complement(560..1375)
/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 1283..1302
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
primer_bind 1367..1386
/label=pENTR-R
/note="pENTR vectors, reverse primer"
rep_origin 1497..2085
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 1986..2005
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 2239..2256
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 2271..2413
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(2339..2358)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 2499..2521
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(2515..2706)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(3515..4597)
/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 4559..4578
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(4598..4675)
/gene="lacI"
/label=lacI promoter
primer_bind 4881..4900
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind 4984..5003
/label=T7
/note="T7 promoter, forward primer"
promoter 4984..5002
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 5003..5027
/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 5042..5064
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 5074..5091
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 5104..5124
/codon_start=1
/product="tobacco etch virus (TEV) protease recognition and
cleavage site"
/label=TEV site
/translation="ENLYFQG"
protein_bind 5152..5185
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
promoter 5200..5645
/gene="sacR"
/label=sacB promoter
/note="sacB promoter and control region"
CDS 5646..7067
/codon_start=1
/gene="Bacillus subtilis sacB"
/product="secreted levansucrase that renders bacterial
growth sensitive to sucrose"
/label=SacB
/note="negative selection marker"
/translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
VKDSILEQGQLTVNK"
CDS 7169..7186
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(7239..7257)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 7253..7300
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
ORIGIN
1 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
61 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
121 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
181 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
241 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
301 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
361 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
421 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaatttcag gtggcacttt
481 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
541 tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat
601 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa
661 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc
721 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga
781 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc
841 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac
901 cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac
961 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat
1021 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag
1081 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca
1141 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac
1201 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg
1261 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca
1321 tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac
1381 cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa
1441 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
1501 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
1561 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
1621 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
1681 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
1741 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
1801 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
1861 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
1921 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
1981 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
2041 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
2101 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
2161 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
2221 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg
2281 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tatatggtgc actctcagta
2341 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg
2401 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
2461 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
2521 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc
2581 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag
2641 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt
2701 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa
2761 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg
2821 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg
2881 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc
2941 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta
3001 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca
3061 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc
3121 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc
3181 catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa
3241 ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc
3301 gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac
3361 gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca
3421 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta
3481 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
3541 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
3601 tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca
3661 ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa
3721 aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt
3781 atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg
3841 cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca
3901 gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta
3961 tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg
4021 agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat
4081 gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct
4141 ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg
4201 catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat
4261 tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc
4321 tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca
4381 gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg
4441 ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt
4501 tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg
4561 catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct
4621 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga
4681 tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg
4741 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc
4801 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg
4861 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg
4921 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatctc gatcccgcga
4981 aattaatacg actcactata ggggaattgt gagcggataa caattcccct ctagaaataa
5041 ttttgtttaa ctttaagaag gagatatacc atgcatcatc atcatcatca cagcagcggc
5101 agagaaaact tgtatttcca gggccatatg agttctcctc ctgaaagatc cataacttcg
5161 tatagcatac attatacgaa gttatgcggc cgcgacgtcc acatatacct gccgttcact
5221 attatttagt gaaatgagat attatgatat tttctgaatt gtgattaaaa aggcaacttt
5281 atgcccatgc aacagaaact ataaaaaata cagagaatga aaagaaacag atagattttt
5341 tagttcttta ggcccgtagt ctgcaaatcc ttttatgatt ttctatcaaa caaaagagga
5401 aaatagacca gttgcaatcc aaacgagagt ctaatagaat gaggtcgaaa agtaaatcgc
5461 gcgggtttgt tactgataaa gcaggcaaga cctaaaatgt gtaaagggca aagtgtatac
5521 tttggcgtca ccccttacat attttaggtc tttttttatt gtgcgtaact aacttgccat
5581 cttcaaacag gagggctgga agaagcagac cgctaacaca gtacataaaa aaggagacat
5641 gaacgatgaa catcaaaaag tttgcaaaac aagcaacagt attaaccttt actaccgcac
5701 tgctggcagg aggcgcaact caagcgtttg cgaaagaaac gaaccaaaag ccatataagg
5761 aaacatacgg catttcccat attacacgcc atgatatgct gcaaatccct gaacagcaaa
5821 aaaatgaaaa atatcaagtt cctgagttcg attcgtccac aattaaaaat atctcttctg
5881 caaaaggcct ggacgtttgg gacagctggc cattacaaaa cgctgacggc actgtcgcaa
5941 actatcacgg ctaccacatc gtctttgcat tagccggaga tcctaaaaat gcggatgaca
6001 catcgattta catgttctat caaaaagtcg gcgaaacttc tattgacagc tggaaaaacg
6061 ctggccgcgt ctttaaagac agcgacaaat tcgatgcaaa tgattctatc ctaaaagacc
6121 aaacacaaga atggtcaggt tcagccacat ttacatctga cggaaaaatc cgtttattct
6181 acactgattt ctccggtaaa cattacggca aacaaacact gacaactgca caagttaacg
6241 tatcagcatc agacagctct ttgaacatca acggtgtaga ggattataaa tcaatctttg
6301 acggtgacgg aaaaacgtat caaaatgtac agcagttcat cgatgaaggc aactacagct
6361 caggcgacaa ccatacgctg agagatcctc actacgtaga agataaaggc cacaaatact
6421 tagtatttga agcaaacact ggaactgaag atggctacca aggcgaagaa tctttattta
6481 acaaagcata ctatggcaaa agcacatcat tcttccgtca agaaagtcaa aaacttctgc
6541 aaagcgataa aaaacgcacg gctgagttag caaacggcgc tctcggtatg attgagctaa
6601 acgatgatta cacactgaaa aaagtgatga aaccgctgat tgcatctaac acagtaacag
6661 atgaaattga acgcgcgaac gtctttaaaa tgaacggcaa atggtacctg ttcactgact
6721 cccgcggatc aaaaatgacg attgacggca ttacgtctaa cgatatttac atgcttggtt
6781 atgtttctaa ttctttaact ggcccataca agccgctgaa caaaactggc cttgtgttaa
6841 aaatggatct tgatcctaac gatgtaacct ttacttactc acacttcgct gtacctcaag
6901 cgaaaggaaa caatgtcgtg attacaagct atatgacaaa cagaggattc tacgcagaca
6961 aacaatcaac gtttgcgcct agcttcctgc tgaacatcaa aggcaagaaa acatctgttg
7021 tcaaagacag catccttgaa caaggacaat taacagttaa caaataaaaa cgcaaaagaa
7081 aatgccgata tcctattggc attgacgtca ggtggcactt ttcgaggaga tcatgcacat
7141 gatgacgaag cttgcggccg cactcgagca ccaccaccac caccactgag atccggctgc
7201 taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata
7261 accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc
7321 cggat
//
