Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V011023 | pET-21a(+) | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pET-21a(+)
- Antibiotic Resistance:
- Ampicillin
- Length:
- 5439 bp
- Type:
- pET & Duet Vectors (Novagen)
- Replication origin:
- ori
- Source/Author:
- Novagen (EMD Millipore)
- Copy Number:
- High copy number
- Growth Strain(s):
- DH10b
- Growth Temperature:
- 37℃
pET-21a(+) 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.
References
- Kim YJ, Shin JS, Lee KW, Eom HJ, Jo BG, Lee JW, Kim JH, Kim SY, Kang JH, Choi JW. Expression, Purification, and Characterization of Plasmodium vivax Lactate Dehydrogenase from Bacteria without Codon Optimization. Int J Mol Sci. 2023 Jul 4;24(13):11083. doi: 10.3390/ijms241311083. PMID: 37446261; PMCID: PMC10342390.
- Zafar A, Aftab MN, Asif A, Karadag A, Peng L, Celebioglu HU, Afzal MS, Hamid A, Iqbal I. Efficient biomass saccharification using a novel cellobiohydrolase from Clostridium clariflavum for utilization in biofuel industry. RSC Adv. 2021 Mar 1;11(16):9246-9261. doi: 10.1039/d1ra00545f. Erratum in: RSC Adv. 2021 Mar 18;11(19):11387. doi: 10.1039/d1ra90090k. PMID: 35423428; PMCID: PMC8695235.
- Wang Y, Xuan G, Lin H, Fei Z, Wang J. Phage resistance of Salmonella enterica obtained by transposon Tn5-mediated SefR gene silent mutation. J Basic Microbiol. 2023 May;63(5):530-541. doi: 10.1002/jobm.202200532. Epub 2023 Apr 9. PMID: 37032321.
pET-21a(+) vector Sequence
LOCUS Exported 5439 bp DNA circular SYN 03-SEP-2024
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pET-21a(+)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5439)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..5439
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 124..142
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 143..167
/label=lac operator
/bound_moiety="lac repressor encoded by 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)."
RBS 182..204
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 212..244
/codon_start=1
/product="leader peptide from bacteriophage T7 gene 10"
/label=T7 tag (gene 10 leader)
/note="promotes efficient translation in E. coli"
/translation="MASMTGGQQMG"
CDS 294..311
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
terminator 378..425
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 462..917
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 943..1047
/gene="bla"
/label=AmpR promoter
CDS 1048..1908
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 2079..2667
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature 2853..2992
/label=bom
/note="basis of mobility region from pBR322"
CDS complement(3094..3285)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(4094..5176)
/codon_start=1
/gene="lacI"
/product="lac repressor"
/label=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"
promoter complement(5177..5254)
/gene="lacI"
/label=lacI promoter