| 质粒名称: | pET28a-SnoopCatcher |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 72322 |
| 存储实验室: | Mark Howarth |
|
载体骨架基本信息 |
|
| 载体名称: | pET28a |
| 载体出品公司: | Novagen |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | 5369 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: | pET28a-SnoopCatcher,SnoopCatcher |
| 物种来源: | Synthetic |
| 插入大小: | 405 bp |
| 基因突变: | 更改肺炎链球菌粘附素RrgA的G842T和D848G（根据PDB ID代码2WW8编号） |
| GenBank编号 | -- |
| 启动子 | T7 |
| 标签/融合蛋白: | His6标签（插入N端子） |
| 克隆方法: | Ligation Independent克隆 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | TAATACGACTCACTATAGGG |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pET28a-SnoopCatcher was a gift from Mark Howarth (Addgene plasmid # 72322 ; http://n2t.net/addgene:72322 ; RRID:Addgene_72322) |
| 参考文献部分: | Programmable polyproteams built using twin peptide superglues. Veggiani G, Nakamura T, Brenner MD, Gayet RV, Yan J, Robinson CV, Howarth M. Proc Natl Acad Sci U S A. 2016 Jan 19. pii: 201519214. 10.1073/pnas.1519214113 PubMed 26787909 |
买家导航
pET28aSnoopCatcher载体基本信息
pET28aSnoopCatcher载体图谱质粒图谱和多克隆位点信息
pET28aSnoopCatcher载体简介
pET28aSnoopCatcher载体序列
LOCUS pET28a-SnoopCatcher 5653 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Bacterial expression of SnoopCatcher, an engineered protein forming
a spontaneous covalent bond with its cognate peptide partner
SnoopTag..
ACCESSION .
VERSION .
KEYWORDS pET28a-SnoopCatcher
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5653)
AUTHORS Veggiani G, Nakamura T, Brenner MD, Gayet RV, Yan J, Robinson CV,
Howarth M
TITLE Programmable polyproteams built using twin peptide superglues.
JOURNAL Proc Natl Acad Sci U S A. 2016 Jan 19. pii: 201519214.
PUBMED 26787909
REFERENCE 2 (bases 1 to 5653)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..5653
/organism="synthetic DNA construct"
/mol_type="other DNA"
terminator 26..73
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
primer_bind 69..87
/label=T7 Term
/note="T7 terminator, reverse primer"
CDS complement(527..544)
/codon_start=1
/product="thrombin recognition and cleavage site"
/label=thrombin site
/translation="LVPRGS"
CDS complement(554..571)
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
RBS 590..612
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
protein_bind 627..651
/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 complement(651..670)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(652..670)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(754..773)
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
promoter 979..1056
/gene="lacI"
/label=lacI promoter
CDS 1057..2139
/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(1076..1095)
/label=LacI-R
/note="LacI, reverse primer"
CDS 2948..3139
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
primer_bind complement(3133..3155)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
misc_feature 3241..3383
/label=bom
/note="basis of mobility region from pBR322"
primer_bind 3296..3315
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind complement(3398..3415)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3569..4157)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3649..3668)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind complement(4268..4287)
/label=pENTR-R
/note="pENTR vectors, reverse primer"
CDS 4279..5094
/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 complement(4352..4371)
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
rep_origin complement(5187..5642)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5324..5345)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 5536..5555
/label=F1ori-R
/note="F1 origin, reverse primer"
ORIGIN
1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
121 tgttagcagc cggatctcag tggtggtggt ggtagtgctc gagtgcggcc gcaagctttt
181 attatttcgg cggtatcggt tcattggtga tataatgttt accgttggta aattcatatg
241 tagccggaat atcctgcggc acaatgctgg tcacatcacg cacttcgcca ttcacaatct
301 gaaacgccac aatcggctta ttctgcaccg gtttatagcc agcgggttcg ctattttcaa
361 acaggcgata tttgccatcg ctcagattct taaaggtcag tttaccatct tcgccggtac
421 gcacattttg ataggtccca ttctgatcaa tcgcgccata gatatcggga tagtcgggat
481 gctgtttctg caggctaaac acggcaccac gcagcggctt catatggctg ccgcgcggca
541 ccaggccgct gctgtgatga tgatgatgat ggctgctgcc catggtatat ctccttctta
601 aagttaaaca aaattatttc tagaggggaa ttgttatccg ctcacaattc ccctatagtg
661 agtcgtatta atttcgcggg atcgagatct cgatcctcta cgccggacgc atcgtggccg
721 gcatcaccgg cgccacaggt gcggttgctg gcgcctatat cgccgacatc accgatgggg
781 aagatcgggc tcgccacttc gggctcatga gcgcttgttt cggcgtgggt atggtggcag
841 gccccgtggc cgggggactg ttgggcgcca tctccttgca tgcaccattc cttgcggcgg
901 cggtgctcaa cggcctcaac ctactactgg gctgcttcct aatgcaggag tcgcataagg
961 gagagcgtcg agatcccgga caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga
1021 tagcgcccgg aagagagtca attcagggtg gtgaatgtga aaccagtaac gttatacgat
1081 gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc
1141 cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt
1201 cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc
1261 tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat
1321 caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa
1381 gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg
1441 gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt
1501 gatgtctctg accagacacc catcaacagt attattttct cccatgaaga cggtacgcga
1561 ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca
1621 ttaagttctg tctcggcgcg tctgcgtctg gctggctggc ataaatatct cactcgcaat
1681 caaattcagc cgatagcgga acgggaaggc gactggagtg ccatgtccgg ttttcaacaa
1741 accatgcaaa tgctgaatga gggcatcgtt cccactgcga tgctggttgc caacgatcag
1801 atggcgctgg gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg tgcggatatc
1861 tcggtagtgg gatacgacga taccgaagac agctcatgtt atatcccgcc gttaaccacc
1921 atcaaacagg attttcgcct gctggggcaa accagcgtgg accgcttgct gcaactctct
1981 cagggccagg cggtgaaggg caatcagctg ttgcccgtct cactggtgaa aagaaaaacc
2041 accctggcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
2101 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag
2161 ttagctcact cattaggcac cgggatctcg accgatgccc ttgagagcct tcaacccagt
2221 cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt
2281 tatcatgcaa ctcgtaggac aggtgccggc agcgctctgg gtcattttcg gcgaggaccg
2341 ctttcgctgg agcgcgacga tgatcggcct gtcgcttgcg gtattcggaa tcttgcacgc
2401 cctcgctcaa gccttcgtca ctggtcccgc caccaaacgt ttcggcgaga agcaggccat
2461 tatcgccggc atggcggccc cacgggtgcg catgatcgtg ctcctgtcgt tgaggacccg
2521 gctaggctgg cggggttgcc ttactggtta gcagaatgaa tcaccgatac gcgagcgaac
2581 gtgaagcgac tgctgctgca aaacgtctgc gacctgagca acaacatgaa tggtcttcgg
2641 tttccgtgtt tcgtaaagtc tggaaacgcg gaagtcagcg ccctgcacca ttatgttccg
2701 gatctgcatc gcaggatgct gctggctacc ctgtggaaca cctacatctg tattaacgaa
2761 gcgctggcat tgaccctgag tgatttttct ctggtcccgc cgcatccata ccgccagttg
2821 tttaccctca caacgttcca gtaaccgggc atgttcatca tcagtaaccc gtatcgtgag
2881 catcctctct cgtttcatcg gtatcattac ccccatgaac agaaatcccc cttacacgga
2941 ggcatcagtg accaaacagg aaaaaaccgc ccttaacatg gcccgcttta tcagaagcca
3001 gacattaacg cttctggaga aactcaacga gctggacgcg gatgaacagg cagacatctg
3061 tgaatcgctt cacgaccacg ctgatgagct ttaccgcagc tgcctcgcgc gtttcggtga
3121 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
3181 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
3241 cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca
3301 tcagagcaga ttgtactgag agtgcaccat atatgcggtg tgaaataccg cacagatgcg
3361 taaggagaaa ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct
3421 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
3481 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
3541 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
3601 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
3661 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
3721 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
3781 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
3841 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
3901 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
3961 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg
4021 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
4081 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
4141 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
4201 acgaaaactc acgttaaggg attttggtca tgaacaataa aactgtctgc ttacataaac
4261 agtaatacaa ggggtgttat gagccatatt caacgggaaa cgtcttgctc taggccgcga
4321 ttaaattcca acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg
4381 caatcaggtg cgacaatcta tcgattgtat gggaagcccg atgcgccaga gttgtttctg
4441 aaacatggca aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg
4501 ctgacggaat ttatgcctct tccgaccatc aagcatttta tccgtactcc tgatgatgca
4561 tggttactca ccactgcgat ccccgggaaa acagcattcc aggtattaga agaatatcct
4621 gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt
4681 cctgtttgta attgtccttt taacagcgat cgcgtatttc gtctcgctca ggcgcaatca
4741 cgaatgaata acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct
4801 gttgaacaag tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc
4861 actcatggtg atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt
4921 attgatgttg gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac
4981 tgcctcggtg agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat
5041 aatcctgata tgaataaatt gcagtttcat ttgatgctcg atgagttttt ctaagaatta
5101 attcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
5161 cacatttccc cgaaaagtgc cacctgaaat tgtaaacgtt aatattttgt taaaattcgc
5221 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
5281 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
5341 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
5401 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
5461 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
5521 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
5581 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
5641 gtcccattcg cca
//
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