Basic Vector Information
- Vector Name:
- pET His6 MBP TEV LIC cloning vector (1M)
- Antibiotic Resistance:
- Kanamycin
- Length:
- 6462 bp
- Type:
- Bacterial Expression
- Replication origin:
- ori
- Copy Number:
- Low Copy
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- MBP forward (5'ggtcgtcagactgtcgatgaagcc)
- 3' Primer:
- T7 reverse
pET His6 MBP TEV LIC cloning vector (1M) vector Vector Map
Plasmid Resuspension Protocol:
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5.Store the plasmid at -20 ℃.
pET His6 MBP TEV LIC cloning vector (1M) vector Sequence
LOCUS 40924_18611 6462 bp DNA circular SYN 13-MAY-2021
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6462)
TITLE LIC N-terminal fusion vectors for E. coli expression
REFERENCE 2 (bases 1 to 6462)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 6462)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6462
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind 106..125
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
promoter 209..227
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 228..252
/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)."
RBS 267..289
/label=RBS
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 308..325
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
CDS 335..1435
/codon_start=1
/label=MBP
/note="maltose binding protein from E. coli"
/translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
WYAVRTAVINAASGRQTVDEALKDAQT"
CDS 1448..1468
/codon_start=1
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
/translation="ENLYFQS"
CDS 1534..1551
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
terminator 1618..1665
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 1702..2157
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
CDS complement(2253..3065)
/codon_start=1
/label=KanR
/note="aminoglycoside phosphotransferase"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
rep_origin 3187..3775
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3929..3946
/label=L4440
/note="L4440 vector, forward primer"
misc_feature complement(3961..4100)
/label=bom
/note="basis of mobility region from pBR322"
primer_bind 4186..4208
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(4205..4393)
/codon_start=1
/label=rop
/note="Rop protein, which maintains plasmids at low copy
number"
/translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
protein_bind complement(5168..5189)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(5205..6284)
/codon_start=1
/label=lacI
/note="lac repressor"
/translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
promoter complement(6285..6362)
/label=lacI promoter
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