质粒名称: | pRK793 |
---|---|
出品公司: | 爱迪基因 |
目录编号: | 8827 |
存储实验室: | David Waugh |
载体骨架基本信息 |
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载体名称: | pMal-C2 |
载体出品公司: | New England Biolabs |
载体抗性: | 氯霉素/氨苄青霉素 |
空载体大小: | 6700 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: | 低拷贝 |
生长条件: | 37 ℃ |
插入基因信息 |
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插入基因名称: | TEV protease, S219V mutant,tobacco etch virus protease |
物种来源: | -- |
插入大小: | 750 bp |
基因突变: | S219V mutation (improves stability) |
GenBank编号 | -- |
启动子 | -- |
标签/融合蛋白: |
His(插入件上的N端子)
polyarginine(插入物上的C端)
MBP(带TEV)(插入件上的N端子)
|
克隆方法: | 限制性内切酶 |
5' 克隆位点: | SacI(未破坏) |
3' 克隆位点: | BamHI (未破坏) |
5' 测序引物 | -- |
3' 测序引物 | M13-F20 |
文献引用方法 |
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材料和方法部分: | pRK793 was a gift from David Waugh (Addgene plasmid # 8827 ; http://n2t.net/addgene:8827 ; RRID:Addgene_8827) |
参考文献部分: | Tobacco etch virus protease: mechanism of autolysis and rational design of stable mutants with wild-type catalytic proficiency. Kapust RB, Tozser J, Fox JD, Anderson DE, Cherry S, Copeland TD, Waugh DS. Protein Eng. 2001 Dec . 14(12):993-1000. 10.1093/protein/14.12.993 PubMed 11809930 |
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LOCUS pRK793 3586 bp ds-DNA circular SYN 06-AUG-2018 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pRK793 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3586) AUTHORS Kapust RB, Tozser J, Fox JD, Anderson DE, Cherry S, Copeland TD, Waugh DS TITLE Tobacco etch virus protease: mechanism of autolysis and rational design of stable mutants with wild-type catalytic proficiency. JOURNAL Protein Eng. 2001 Dec . 14(12):993-1000. PUBMED 11809930 REFERENCE 2 (bases 1 to 3586) AUTHORS . TITLE Direct Submission JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3586 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 97..114 /label=L4440 /note="L4440 vector, forward primer" promoter 506..608 /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" CDS 609..1268 /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind complement(675..694) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" tRNA complement(2561..2637) /gene="argU" /product="E. coli tRNA-Arg recognizing AGA and AGG" /label=argU promoter complement(2881..2909) /label=tet promoter /note="E. coli promoter for tetracycline efflux protein gene" rep_origin 3021..3566 /direction=RIGHT /label=p15A ori /note="Plasmids containing the medium-copy-number p15A origin of replication can be propagated in E. coli cells that contain a second plasmid with the ColE1 origin. " primer_bind 3467..3486 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 ctcacttccc tgttaagtat cttcctggca tcttccagga aatctccgcc ccgttcgtaa 61 gccatttccg ctcgccgcag tcgaacgacc gagcgtagcg agtcagtgag cgaggaagcg 121 gaatatatcc tgtatcacat attctgctga cgcaccggtg cagccttttt tctcctgcca 181 catgaagcac ttcactgaca ccctcatcag tgccaacata gtaagccagt atacactccg 241 ctagcgctga tgtccggcgg tgcttttgcc gttacgcacc accccgtcag tagctgaaca 301 ggagggacag ctgatagaaa cagaagccac tggagcacct caaaaacacc atcatacact 361 aaatcagtaa gttggcagca tcacccgacg cactttgcgc cgaataaata cctgtgacgg 421 aagatcactt cgcagaataa ataaatcctg gtgtccctgt tgataccggg aagccctggg 481 ccaacttttg gcgaaaatga gacgttgatc ggcacgtaag aggttccaac tttcaccata 541 atgaaataag atcactaccg ggcgtatttt ttgagttatc gagattttca ggagctaagg 601 aagctaaaat ggagaaaaaa atcactggat ataccaccgt tgatatatcc caatggcatc 661 gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg tacctataac cagaccgttc 721 agctggatat tacggccttt ttaaagaccg taaagaaaaa taagcacaag ttttatccgg 781 cctttattca cattcttgcc cgcctgatga atgctcatcc ggaattccgt atggcaatga 841 aagacggtga gctggtgata tgggatagtg ttcacccttg ttacaccgtt ttccatgagc 901 aaactgaaac gttttcatcg ctctggagtg aataccacga cgatttccgg cagtttctac 961 acatatattc gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc cctaaagggt 1021 ttattgagaa tatgtttttc gtctcagcca atccctgggt gagtttcacc agttttgatt 1081 taaacgtggc caatatggac aacttcttcg cccccgtttt caccatgggc aaatattata 1141 cgcaaggcga caaggtgctg atgccgctgg cgattcaggt tcatcatgcc gtttgtgatg 1201 gcttccatgt cggcagaatg cttaatgaat tacaacagta ctgcgatgag tggcagggcg 1261 gggcgtaatt tttttaaggc agttattggt gcccttaaac gcctggttgc tacgcctgaa 1321 taagtgataa taagcggatg aatggcagaa attcgaaagc aaattcgacc cggtcgtcgg 1381 ttcagggcag ggtcgttaaa tagccgctta tgtctattgc tggtttaccg gtttattgac 1441 taccggaagc agtgtgaccg tgtgcttctc aaatgcctga ggccagtttg ctcaggctct 1501 ccccgtggag gtaataattg acgatatgat catttattct gcctcccaga gcctgataaa 1561 aacggttagc gcttcgttaa tacagatgta ggtgttccac agggtagcca gcagcatcct 1621 gcgatgcaga tccggaacat aatggtgcag ggcgcttgtt tcggcgtggg tatggtggca 1681 ggccccgtgg ccgggggact gttgggcgct gccggcacct gtcctacgag ttgcatgata 1741 aagaagacag tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg gaaggagcta 1801 ccggcagcgg tgcggactgt tgtaactcag aataagaaat gaggccgctc atggcgttga 1861 ctctcagtca tagtatcgtg gtatcaccgg ttggttccac tctctgttgc gggcaacttc 1921 agcagcacgt aggggacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca 1981 ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc 2041 gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca 2101 acgacaggag cacgatcatg cgcacccgtg gccaggaccc aacgctgccc gactcgaggg 2161 catccgatca acgctttctg gtgaatggtg cgggaggcga gacttgaact cgcacaccta 2221 gcggcgccag aacctaaatc tggtgcgtct accaatttcg ccactcccgc aaaaaaagat 2281 ggtggctacg acgggattct ctagatcatc atgtttattg cgtggtttat aagtattgtt 2341 attgcacact caaggccctt tagctcagtg gttagagcag gcgactcata atcgcttggt 2401 cgctggttca agtccagcaa gggccaccaa atcattattt acgtatcagg aagagttaag 2461 ggtacaaggg gtactcgatg gcggtctata tttcatgact aacgccattt caaggccatg 2521 gctggaggat ataaagaagg cgtaactgat tgaattgtaa tggcgcgccc tgcaggattc 2581 gaacctgcgg cccacgactt agaaggtcgt tgctctatcc aactgagcta agggcgcgtt 2641 gataccgcaa tgcggtgtaa tcgcgtgaat tatacggtca acccttgctg agtcaatggc 2701 ttttgatctg gttgctgaac aagtgaacga ccgcgtctga ttactagttt gcccgggccc 2761 ttgtcgtcgt caattgtaca gcatccaggg tgacggtgcc gaggatgacg atgagcgcat 2821 tgttagattt catacacggt gcctgactgc gttagcaatt taactgtgat aaactaccgc 2881 attaaagctt atcgatgata agctgtcaaa catgagaatt acaacttata tcgtatgggg 2941 ctgacttcag gtgctacatt tgaagagata aattgcactg aaatctagaa atattttatc 3001 tgattaataa gatgatcttc ttgagatcgt tttggtctgc gcgtaatctc ttgctctgaa 3061 aacgaaaaaa ccgccttgca gggcggtttt tcgaaggttc tctgagctac caactctttg 3121 aaccgaggta actggcttgg aggagcgcag tcaccaaaac ttgtcctttc agtttagcct 3181 taaccggcgc atgacttcaa gactaactcc tctaaatcaa ttaccagtgg ctgctgccag 3241 tggtgctttt gcatgtcttt ccgggttgga ctcaagacga tagttaccgg ataaggcgca 3301 gcggtcggac tgaacggggg gttcgtgcat acagtccagc ttggagcgaa ctgcctaccc 3361 ggaactgagt gtcaggcgtg gaatgagaca aacgcggcca taacagcgga atgacaccgg 3421 taaaccgaaa ggcaggaaca ggagagcgca cgagggagcc gccaggggga aacgcctggt 3481 atctttatag tcctgtcggg tttcgccacc actgatttga gcgtcagatt tcgtgatgct 3541 tgtcaggggg gcggagccta tggaaaaacg gctttgccgc ggccct //
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