载体基本信息 | |||
载体名称 | pMCSG29 | 载体抗性 | Ampicillin |
---|---|---|---|
载体长度 | 6389 bp | 载体类型 | Structural Genomics Vectors |
载体来源 | Eschenfeldt WH, Maltseva N, Stols L, Donnelly MI, Gu M, Nocek B, Tan | 拷贝数 | High copy number |
pMCSG29 载体载体图谱
pMCSG29 载体载体序列
LOCUS Exported 6389 bp ds-DNA circular SYN 11-2-2014 DEFINITION Bacterial vector encoding a C-terminal TEV-10xHis-TVMV-MBP cassette, for high-throughput purification of recombinant proteins. ACCESSION . VERSION . KEYWORDS pMCSG29 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6389) 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 6389) AUTHORS Midwest Center for Structural Genomics TITLE Direct Submission COMMENT For ligation-independent cloning (LIC), linearize with SmaI and treat with T4 DNA polymerase plus dATP. FEATURES Location/Qualifiers source 1..6389 /organism="synthetic DNA construct" /lab_host="Escherichia coli" /mol_type="other DNA" terminator 26..73 /note="T7 terminator" /note="transcription terminator for bacteriophage T7 RNA polymerase" CDS complement(140..157) /codon_start=1 /product="6xHis affinity tag" /note="6xHis" /translation="HHHHHH" CDS complement(213..1310) /codon_start=1 /gene="malE (mutated)" /product="maltose binding protein from E. coli" /note="MBP" /note="This version of the gene does not encode a signal sequence, so MBP will remain in the cytosol." /translation="KIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE NYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFW YAVRTAVINAASGRQTVDEALKDAQT" CDS complement(1311..1331) /codon_start=1 /product="tobacco vein mottling virus (TVMV) NIa protease recognition and cleavage site" /note="TVMV site" /translation="ETVRFQS" CDS complement(1332..1361) /codon_start=1 /product="10xHis affinity tag" /note="10xHis" /translation="HHHHHHHHHH" CDS complement(1368..1388) /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /note="TEV site" /translation="ENLYFQS" RBS 1400..1405 /note="ribosome binding site" protein_bind 1436..1460 /bound_moiety="lac repressor encoded by lacI" /note="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(1461..1479) /note="T7 promoter" /note="promoter for bacteriophage T7 RNA polymerase" promoter 1792..1869 /gene="lacI" /note="lacI promoter" /note=" " CDS 1870..2952 /codon_start=1 /gene="lacI" /product="lac repressor" /note="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" CDS 3761..3952 /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /note="rop" /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" rep_origin complement(4382..4970) /direction=LEFT /note="ori" /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(5141..6001) /codon_start=1 /gene="bla" /product="beta-lactamase" /note="AmpR" /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6002..6105) /gene="bla" /note="AmpR promoter"
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