Basic Vector Information
- Vector Name:
- pMCSG32
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6418 bp
- Type:
- Structural Genomics Vectors
- Replication origin:
- ori
- Source/Author:
- Eschenfeldt WH, Maltseva N, Stols L, Donnelly MI, Gu M, Nocek B, Tan
- Copy Number:
- High copy number
pMCSG32 vector Map
pMCSG32 vector Sequence
LOCUS 40924_30161 6418 bp DNA circular SYN 01-JAN-1980 DEFINITION Bacterial vector encoding N-terminal MBP-TVMV and C-terminal TEV-6xHis, for high-throughput purification of recombinant proteins. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6418) AUTHORS Eschenfeldt WH, Maltseva N, Stols L, Donnelly MI, Gu M, Nocek B, Tan K, Kim Y, Joachimiak A. TITLE Cleavable C-terminal His-tag vectors for structure determination. JOURNAL J. Struct. Funct. Genomics 2010;11:31-9. PUBMED 20213425 REFERENCE 2 (bases 1 to 6418) AUTHORS Midwest Center for Structural Genomics TITLE Direct Submission REFERENCE 3 (bases 1 to 6418) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "J. Struct. Funct. Genomics"; date: "2010"; volume: "11"; pages: "31-9" COMMENT SGRef: number: 2; type: "Journal Article" COMMENT For ligation-independent cloning (LIC), linearize with SmaI and treat with T4 DNA polymerase plus dATP. FEATURES Location/Qualifiers source 1..6418 /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(208..225) /codon_start=1 /label=6xHis /note="6xHis affinity tag" /translation="HHHHHH" CDS complement(232..252) /codon_start=1 /label=TEV site /note="tobacco etch virus (TEV) protease recognition and cleavage site" /translation="ENLYFQS" CDS complement(263..283) /codon_start=1 /label=TVMV site /note="tobacco vein mottling virus (TVMV) NIa protease recognition and cleavage site" /translation="ETVRFQS" protein_bind complement(286..310) /label=attB1 /note="recombination site for the Gateway(R) BP reaction" CDS complement(320..1420) /codon_start=1 /label=MBP /note="maltose binding protein from E. coli" /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF WYAVRTAVINAASGRQTVDEALKDAQT" RBS complement(1428..1450) /label=RBS /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" protein_bind complement(1465..1489) /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(1490..1508) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" promoter 1821..1898 /label=lacI promoter CDS 1899..2978 /codon_start=1 /label=lacI /note="lac repressor" /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR ALADSLMQLARQVSRLESGQ" protein_bind 2994..3015 /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." CDS 3790..3978 /codon_start=1 /label=rop /note="Rop protein, which maintains plasmids at low copy number" /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" misc_feature 4083..4225 /label=bom /note="basis of mobility region from pBR322" rep_origin complement(4411..4999) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(5173..6030) /codon_start=1 /label=AmpR /note="beta-lactamase" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6031..6134) /label=AmpR promoter
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