pCas12f1 vector (V017220)

Price Information

Cat No. Plasmid Name Availability Add to cart
V017220 pCas12f1 In stock, 1 week for quality controls

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Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

Vector Name:
pCas12f1
Antibiotic Resistance:
Kanamycin
Length:
10163 bp
Type:
Genome editing
Replication origin:
pSC101 ori
Host:
E. coli
Promoter:
araBAD
Growth Temperature:
30℃

pCas12f1 vector Vector Map

pCas12f110163 bp50010001500200025003000350040004500500055006000650070007500800085009000950010000lacIlacIq promoterRep101(Ts)pSC101 oriKanRtracrRNACRISPR-associated endodeoxyribonuclease Cas12f1araCaraBAD promoterGamBetaExo

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.

pCas12f1 vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       V017220                10163 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V017220
VERSION     V017220
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 10163)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..10163
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(533..1612)
                     /label="lacI"
                     /note="lac repressor"
     promoter        complement(1613..1690)
                     /label="lacIq promoter"
                     /note="In the lacIq allele, a single base change in the
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             complement(1826..2773)
                     /label="Rep101(Ts)"
                     /note="temperature-sensitive version of the RepA protein
                     needed for replication with the pSC101 origin (Armstrong et
                     al., 1984)"
     rep_origin      complement(2821..3043)
                     /direction=LEFT
                     /label="pSC101 ori"
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             complement(3698..4510)
                     /label="KanR"
                     /note="aminoglycoside phosphotransferase"
     misc_RNA        complement(4666..4744)
                     /label="tracrRNA"
                     /note="trans-activating CRISPR RNA for the Streptococcus
                     pyogenes CRISPR/Cas9 system"
     CDS             5046..6632
                     /gene="cas12f"
                     /label="CRISPR-associated endodeoxyribonuclease Cas12f1"
                     /note="CRISPR-associated endodeoxyribonuclease Cas12f1 from
                     Uncultured archaeon. Accession#: A0A482D308"
     CDS             complement(6774..7649)
                     /label="araC"
                     /note="L-arabinose regulatory protein"
     promoter        7676..7960
                     /label="araBAD promoter"
                     /note="promoter of the L-arabinose operon of E. coli; the
                     araC regulatory gene is transcribed in the opposite
                     direction (Guzman et al., 1995)"
     CDS             8004..8417
                     /label="Gam"
                     /note="inhibitor of the host RecBCD nuclease in the lambda
                     Red system"
     CDS             8426..9208
                     /label="Beta"
                     /note="single-stranded DNA binding recombinase in the
                     lambda Red system"
     CDS             9208..9885
                     /label="Exo"
                     /note="5' to 3' double-stranded DNA exonuclease in the
                     lambda Red system"