Price Information
Cat No. | Plasmid Name | Availability | Add to cart |
---|---|---|---|
V011033 | pET-11b | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pET-11b
- Antibiotic Resistance:
- Ampicillin
- Length:
- 5676 bp
- Type:
- pET & Duet Vectors (Novagen)
- Replication origin:
- ori
- Source/Author:
- Novagen (EMD Millipore)
- Copy Number:
- High copy number
pET-11b vector Map
Plasmid 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 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
pET-11b vector Sequence
LOCUS 40924_17636 5676 bp DNA circular SYN 18-DEC-2018 DEFINITION Bacterial expression vector pET-11b, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5676) AUTHORS De Schamphelaire W, Olbrechts A, Meert J, Verhelst K, Roggeman Fonseca M, Vanhoucke M, Beyaert R. TITLE BCCM/LMBP Plasmid collection JOURNAL Unpublished REFERENCE 2 (bases 1 to 5676) AUTHORS De Schamphelaire W. TITLE Direct Submission JOURNAL Submitted (02-FEB-2017) BCCM/LMBP, Universiteit Gent, Technologiepark 927, 9052, BELGIUM REFERENCE 3 (bases 1 to 5676) TITLE Direct Submission REFERENCE 4 (bases 1 to 5676) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Unpublished" COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (02-FEB-2017) BCCM/LMBP, Universiteit Gent, Technologiepark 927, 9052, BELGIUM" COMMENT SGRef: number: 3; type: "Journal Article" FEATURES Location/Qualifiers source 1..5676 /mol_type="other DNA" /organism="synthetic DNA construct" promoter 10..38 /label=tet promoter /note="E. coli promoter for tetracycline efflux protein gene" terminator complement(213..260) /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" CDS complement(327..359) /codon_start=1 /label=T7 tag (gene 10 leader) /note="leader peptide from bacteriophage T7 gene 10" /translation="MASMTGGQQMG" RBS complement(367..389) /label=RBS /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" protein_bind complement(404..428) /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(429..447) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" promoter 756..833 /label=lacI promoter CDS 834..1913 /codon_start=1 /label=lacI /note="lac repressor" /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR ALADSLMQLARQVSRLESGQ" protein_bind 1929..1950 /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." CDS 3228..3416 /codon_start=1 /label=rop /note="Rop protein, which maintains plasmids at low copy number" /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" misc_feature 3521..3663 /label=bom /note="basis of mobility region from pBR322" rep_origin complement(3849..4437) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(4611..5468) /codon_start=1 /label=AmpR /note="beta-lactamase" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(5469..5573) /label=AmpR promoter