Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V018106 | pKNOCK-Gm | In stock, 1 week for quality controls |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
low copy number in pir+ E. coli strains high copy number in pir116 E. coli strains
- Vector Name:
- pKNOCK-Gm
- Antibiotic Resistance:
- Gentamycin
- Length:
- 1698 bp
- Type:
- Basic Cloning Vectors
- Source/Author:
- Mikhail Alexeyev
- Copy Number:
- Low copy number
- Growth Strain(s):
- DH5a-λpir
- Growth Temperature:
- 37℃
pKNOCK-Gm 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.
pKNOCK-Gm vector Sequence
LOCUS Exported 1698 bp DNA circular SYN 02-JUL-2025
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 1698)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..1698
/mol_type="other DNA"
/organism="synthetic DNA construct"
rep_origin 33..421
/label=R6K gamma ori
/note="gamma replication origin from E. coli plasmid R6K;
requires the R6K initiator protein pi for replication"
primer_bind 454..470
/label=SK primer
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(504..520)
/label=KS primer
/note="common sequencing primer, one of multiple similar
variants"
promoter 559..587
/gene="intI1 (promoter lies within the coding sequence)"
/label=Pc promoter
/note="class 1 integron promoter"
CDS 776..1309
/codon_start=1
/gene="aacC1"
/product="gentamycin acetyltransferase"
/label=GmR
/note="confers resistance to gentamycin"
/translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
EEVMHFDIDPSTAT"
oriT 1451..1560
/note="incP origin of transfer"