载体基本信息 | |||
载体名称 | pMCSG19 | 载体抗性 | Ampicillin |
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载体长度 | 6441 bp | 载体类型 | Structural Genomics Vectors |
载体来源 | Donnelly MI, Zhou M, Millard CS, Clancy S, Stols L, Eschenfeldt WH, | 拷贝数 | High copy number |
pMCSG19 载体载体图谱
pMCSG19 载体载体序列
LOCUS 40924_30086 6441 bp DNA circular SYN 01-JAN-1980 DEFINITION Bacterial vector with an MBP-TVMV-6xHis-TEV leader for high-throughput purification of recombinant proteins. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6441) AUTHORS Donnelly MI, Zhou M, Millard CS, Clancy S, Stols L, Eschenfeldt WH, Collart FR, Joachimiak A. TITLE An expression vector tailored for large-scale, high-throughput purification of recombinant proteins. JOURNAL Protein Expr. Purif. 2006;47:446-54. PUBMED 16497515 REFERENCE 2 (bases 1 to 6441) AUTHORS Midwest Center for Structural Genomics TITLE Direct Submission REFERENCE 3 (bases 1 to 6441) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Protein Expr. Purif."; date: "2006"; volume: "47"; pages: "446-54" COMMENT SGRef: number: 2; type: "Journal Article" COMMENT For ligation-independent cloning (LIC), linearize with SspI and treat with T4 DNA polymerase plus dGTP. FEATURES Location/Qualifiers source 1..6441 /mol_type="other DNA" /organism="synthetic DNA construct" terminator complement(26..73) /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" CDS complement(140..157) /codon_start=1 /label=6xHis /note="6xHis affinity tag" /translation="HHHHHH" CDS complement(223..243) /codon_start=1 /label=TEV site /note="tobacco etch virus (TEV) protease recognition and cleavage site" /translation="ENLYFQS" CDS complement(268..285) /codon_start=1 /label=6xHis /note="6xHis affinity tag" /translation="HHHHHH" CDS complement(286..306) /codon_start=1 /label=TVMV site /note="tobacco vein mottling virus (TVMV) NIa protease recognition and cleavage site" /translation="ETVRFQS" protein_bind complement(309..333) /label=attB1 /note="recombination site for the Gateway(R) BP reaction" CDS complement(343..1443) /codon_start=1 /label=MBP /note="maltose binding protein from E. coli" /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF WYAVRTAVINAASGRQTVDEALKDAQT" RBS complement(1451..1473) /label=RBS /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" protein_bind complement(1488..1512) /label=lac operator /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)." promoter complement(1513..1531) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" promoter 1844..1921 /label=lacI promoter CDS 1922..3001 /codon_start=1 /label=lacI /note="lac repressor" /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR ALADSLMQLARQVSRLESGQ" protein_bind 3017..3038 /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." CDS 3813..4001 /codon_start=1 /label=rop /note="Rop protein, which maintains plasmids at low copy number" /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" misc_feature 4106..4248 /label=bom /note="basis of mobility region from pBR322" rep_origin complement(4434..5022) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(5196..6053) /codon_start=1 /label=AmpR /note="beta-lactamase" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6054..6157) /label=AmpR promoter
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