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pCMV-AsCas12f1 vector (V017094)

Price Information

Cat No. Plasmid Name Availability Add to cart
V017094 pCMV-AsCas12f1 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:
pCMV-AsCas12f1
Antibiotic Resistance:
Ampicillin
Length:
6845 bp
Type:
Gene knockout
Replication origin:
ori
Host:
Mammalian cells
Selection Marker:
Puro
Promoter:
CMV
Growth Strain(s):
DH5a
Growth Temperature:
37℃

pCMV-AsCas12f1 vector Map

Copy Sequence View Map
pCMV-AsCas12f16845 bp3006009001200150018002100240027003000330036003900420045004800510054005700600063006600CMV enhancerCMV promoterT7 promoterSV40 NLSCRISPR-associated endodeoxyribonuclease Cas12f1SV40 NLS6xHisbGH poly(A) signalM13 revAmpR promoterAmpRoriSV40 poly(A) signalPuroRhPGK promotercPPT/CTSU6 promoterpause site

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.

pCMV-AsCas12f1 vector Sequence

LOCUS       V017094                 6845 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V017094
VERSION     V017094
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 6845)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6845
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     promoter        639..657
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             677..697
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             698..1963
                     /gene="cas12f1"
                     /label="CRISPR-associated endodeoxyribonuclease Cas12f1"
                     /note="CRISPR-associated endodeoxyribonuclease Cas12f1 from
                     Sulfoacidibacillus thermotolerans. Accession#: A0A2U3D0N8"
     CDS             1976..1996
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             2005..2022
                     /label="6xHis"
                     /note="6xHis affinity tag"
     polyA_signal    2051..2275
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2346..2362)
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     promoter        2389..2493
                     /label="AmpR promoter"
     CDS             2494..3351
                     /label="AmpR"
                     /note="beta-lactamase"
     rep_origin      3525..4113
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     polyA_signal    4532..4653
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     CDS             complement(4803..5399)
                     /label="PuroR"
                     /note="puromycin N-acetyltransferase"
     promoter        complement(5421..5931)
                     /label="hPGK promoter"
                     /note="human phosphoglycerate kinase 1 promoter"
     misc_feature    complement(5980..6097)
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     promoter        complement(6352..6592)
                     /label="U6 promoter"
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_feature    complement(6671..6762)
                     /label="pause site"
                     /note="RNA polymerase II transcriptional pause signal from
                     the human alpha-2 globin gene"