| 质粒名称: | pET28GST-LIC |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 26101 |
| 存储实验室: | Cheryl Arrowsmith |
|
载体骨架基本信息 |
|
| 载体名称: | pET28GST-LIC |
| 载体出品公司: | Structural Genomics Consortium |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | 7998 bp |
| 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 | -- |
| 标签/融合蛋白: |
GST（主干上的N终端）
6XHis（主干网上的N个终端）
凝血酶裂解位点（主干N端）
|
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | T7, pGEX 5' |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pET28GST-LIC was a gift from Cheryl Arrowsmith (Addgene plasmid # 26101 ; http://n2t.net/addgene:26101 ; RRID:Addgene_26101) |
买家导航
pET28GSTLIC载体基本信息
pET28GSTLIC载体图谱质粒图谱和多克隆位点信息
pET28GSTLIC载体简介
pET28GSTLIC载体序列
LOCUS pET28GST-LIC 7994 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION SGC Empty backbone for bacterial expression.
ACCESSION .
VERSION .
KEYWORDS pET28GST-LIC
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7994)
TITLE Cloning Vectors
REFERENCE 2 (bases 1 to 7994)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7994
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_feature 5..144
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(70..89)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 230..252
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(246..437)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(1246..2328)
/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 2290..2309
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(2329..2406)
/gene="lacI"
/label=lacI promoter
primer_bind 2612..2631
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind 2715..2734
/label=T7
/note="T7 promoter, forward primer"
promoter 2715..2733
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 2734..2758
/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 2773..2795
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 2803..3456
/codon_start=1
/product="glutathione S-transferase from Schistosoma
japonicum"
/label=GST
/translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
primer_bind 3414..3436
/label=pGEX 5'
/note="pGEX vectors, Glutathione-S-transferase, forward
primer"
CDS 3484..3501
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 3511..3528
/codon_start=1
/product="thrombin recognition and cleavage site"
/label=thrombin site
/translation="LVPRGS"
protein_bind 3556..3589
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
promoter 3604..4049
/gene="sacR"
/label=sacB promoter
/note="sacB promoter and control region"
CDS 4050..5471
/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
ITRHDMLQIPEQQKNEKYKVPEFDSSTIKNISSAKGLDVWDSWPLQNTDGTVANYHGYH
IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
VKDSILEQGQLTVNK"
CDS 5573..5590
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(5643..5661)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 5657..5704
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 5741..6196
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5828..5847)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 6038..6059
/label=F1ori-F
/note="F1 origin, forward primer"
CDS complement(6288..7103)
/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 7011..7030
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
primer_bind 7095..7114
/label=pENTR-R
/note="pENTR vectors, reverse primer"
rep_origin 7225..7813
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 7714..7733
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 7967..7984
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcaatggtg
61 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg
121 ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga
181 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
241 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca
301 tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg
361 agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt
421 ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg
481 ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga acatgcccgg
541 ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga ccagagaaaa
601 atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc acagggtagc
661 cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga cttccgcgtt
721 tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac
781 gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca
841 gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgcgcacc
901 cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga
961 ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg
1021 atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc
1081 acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg
1141 ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat
1201 cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc
1261 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
1321 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
1381 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
1441 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
1501 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
1561 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
1621 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
1681 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
1741 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
1801 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
1861 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
1921 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
1981 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
2041 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
2101 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
2161 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
2221 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
2281 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
2341 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
2401 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
2461 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc
2521 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
2581 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
2641 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct
2701 cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata acaattcccc
2761 tctagaaata attttgttta actttaagaa ggagatatac atatgtcccc tatactaggt
2821 tattggaaaa ttaagggcct tgtgcaaccc actcgacttc ttttggaata tcttgaagaa
2881 aaatatgaag agcatttgta tgagcgcgat gaaggtgata aatggcgaaa caaaaagttt
2941 gaattgggtt tggagtttcc caatcttcct tattatattg atggtgatgt taaattaaca
3001 cagtctatgg ccatcatacg ttatatagct gacaagcaca acatgttggg tggttgtcca
3061 aaagagcgtg cagagatttc aatgcttgaa ggagcggttt tggatattag atacggtgtt
3121 tcgagaattg catatagtaa agactttgaa actctcaaag ttgattttct tagcaagcta
3181 cctgaaatgc tgaaaatgtt cgaagatcgt ttatgtcata aaacatattt aaatggtgat
3241 catgtaaccc atcctgactt catgttgtat gacgctcttg atgttgtttt atacatggac
3301 ccaatgtgcc tggatgcgtt cccaaaatta gtttgtttta aaaaacgtat tgaagctatc
3361 ccacaaattg ataagtactt gaaatccagc aagtatatag catggccttt gcagggctgg
3421 caagccacgt ttggtggtgg cgaccatcct ccaaaatcgg atggttcatc catgggcagc
3481 agccatcatc atcatcatca cagcagcggc ctggttccgc gtggtagtat tatgagttct
3541 cctcctgaaa gatccataac ttcgtatagc atacattata cgaagttatg cggccgcgac
3601 gtccacatat acctgccgtt cactattatt tagtgaaatg agatattatg atattttctg
3661 aattgtgatt aaaaaggcaa ctttatgccc atgcaacaga aactataaaa aatacagaga
3721 atgaaaagaa acagatagat tttttagttc tttaggcccg tagtctgcaa atccttttat
3781 gattttctat caaacaaaag aggaaaatag accagttgca atccaaacga gagtctaata
3841 gaatgaggtc gaaaagtaaa tcgcgcgggt ttgttactga taaagcaggc aagacctaaa
3901 atgtgtaaag ggcaaagtgt atactttggc gtcacccctt acatatttta ggtctttttt
3961 tattgtgcgt aactaacttg ccatcttcaa acaggagggc tggaagaagc agaccgctaa
4021 cacagtacat aaaaaaggag acatgaacga tgaacatcaa aaagtttgca aaacaagcaa
4081 cagtattaac ctttactacc gcactgctgg caggaggcgc aactcaagcg tttgcgaaag
4141 aaacgaacca aaagccatat aaggaaacat acggcatttc ccatattaca cgccatgata
4201 tgctgcaaat ccctgaacag caaaaaaatg aaaaatataa agttcctgag ttcgattcgt
4261 ccacaattaa aaatatctct tctgcaaaag gcctggacgt ttgggacagc tggccattac
4321 aaaacactga cggcactgtc gcaaactatc acggctacca catcgtcttt gcattagccg
4381 gagatcctaa aaatgcggat gacacatcga tttacatgtt ctatcaaaaa gtcggcgaaa
4441 cttctattga cagctggaaa aacgctggcc gcgtctttaa agacagcgac aaattcgatg
4501 caaatgattc tatcctaaaa gaccaaacac aagaatggtc aggttcagcc acatttacat
4561 ctgacggaaa aatccgttta ttctacactg atttctccgg taaacattac ggcaaacaaa
4621 cactgacaac tgcacaagtt aacgtatcag catcagacag ctctttgaac atcaacggtg
4681 tagaggatta taaatcaatc tttgacggtg acggaaaaac gtatcaaaat gtacagcagt
4741 tcatcgatga aggcaactac agctcaggcg acaaccatac gctgagagat cctcactacg
4801 tagaagataa aggccacaaa tacttagtat ttgaagcaaa cactggaact gaagatggct
4861 accaaggcga agaatcttta tttaacaaag catactatgg caaaagcaca tcattcttcc
4921 gtcaagaaag tcaaaaactt ctgcaaagcg ataaaaaacg cacggctgag ttagcaaacg
4981 gcgctctcgg tatgattgag ctaaacgatg attacacact gaaaaaagtg atgaaaccgc
5041 tgattgcatc taacacagta acagatgaaa ttgaacgcgc gaacgtcttt aaaatgaacg
5101 gcaaatggta cctgttcact gactcccgcg gatcaaaaat gacgattgac ggcattacgt
5161 ctaacgatat ttacatgctt ggttatgttt ctaattcttt aactggccca tacaagccgc
5221 tgaacaaaac tggccttgtg ttaaaaatgg atcttgatcc taacgatgta acctttactt
5281 actcacactt cgctgtacct caagcgaaag gaaacaatgt cgtgattaca agctatatga
5341 caaacagagg attctacgca gacaaacaat caacgtttgc gcctagcttc ctgctgaaca
5401 tcaaaggcaa gaaaacatct gttgtcaaag acagcatcct tgaacaagga caattaacag
5461 ttaacaaata aaaacgcaaa agaaaatgcc gatatcctat tggcattgac gtcaggtggc
5521 acttttcgag gagatcatgc acatgatgac gaagcttgcg gccgcactcg agcaccacca
5581 ccaccaccac tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc
5641 caccgctgag caataactag cataacccct tggggcctct aaacgggtct tgaggggttt
5701 tttgctgaaa ggaggaacta tatccggatt ggcgaatggg acgcgccctg tagcggcgca
5761 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
5821 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
5881 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
5941 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
6001 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
6061 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
6121 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
6181 ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
6241 tatttttcta aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca
6301 tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga
6361 aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga
6421 tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc
6481 tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag
6541 aatggcaaaa gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg
6601 tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga
6661 cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc
6721 aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc
6781 tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg
6841 ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc
6901 tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca
6961 tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc
7021 catttatacc catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac
7081 gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt
7141 tttattgttc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
7201 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
7261 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
7321 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt
7381 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
7441 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
7501 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
7561 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
7621 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
7681 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
7741 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
7801 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
7861 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
7921 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
7981 cgaggaagcg gaag
//
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pET28GSTLIC载体质粒应用举例
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