Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V007094 | pLenti SpBsmBI sgRNA Puro | 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:
- pLenti SpBsmBI sgRNA Puro
- Antibiotic Resistance:
- Ampicillin
- Length:
- 8475 bp
- Type:
- Mammalian Expression, Lentiviral
- Replication origin:
- ori
- Selection Marker:
- Puromycin
- Promoter:
- EF-1α
pLenti SpBsmBI sgRNA Puro 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.
pLenti SpBsmBI sgRNA Puro vector Sequence
LOCUS V007094 8475 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V007094
VERSION V007094
KEYWORDS pLenti SpBsmBI sgRNA Puro
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 8475)
AUTHORS Pham H, Kearns NA, Maehr R
TITLE Transcriptional Regulation with CRISPR/Cas9 Effectors in Mammalian
Cells.
JOURNAL Methods Mol Biol. 2016;1358:43-57. doi: 10.1007/978-1-4939-3067-8_3.
PUBMED 26463376
REFERENCE 2 (bases 1 to 8475)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 8475)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Methods Mol
Biol."; date: "2016"; pages: "
10.1007/978-1-4939-3067-8_3"
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..8475
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter complement(20..38)
/label="T3 promoter"
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(59..75)
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(83..99)
/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(107..137)
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind complement(152..173)
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(290..307)
/label="L4440"
/note="L4440 vector, forward primer"
rep_origin complement(461..1049)
/direction=LEFT
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(1223..2080)
/label="AmpR"
/note="beta-lactamase"
promoter complement(2081..2185)
/label="AmpR promoter"
rep_origin complement(2211..2666)
/direction=LEFT
/label="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind 2808..2824
/label="M13 fwd"
/note="common sequencing primer, one of multiple similar
variants"
promoter 2834..2852
/label="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
rep_origin complement(2873..3008)
/direction=LEFT
/label="SV40 ori"
/note="SV40 origin of replication"
polyA_signal complement(3035..3169)
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
LTR complement(3241..3474)
/label="3' LTR (Delta-U3)"
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
misc_feature complement(3561..4149)
/label="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
CDS complement(4159..4755)
/label="PuroR"
/note="puromycin N-acetyltransferase"
promoter complement(4778..5959)
/label="EF-1-alpha promoter"
/note="strong constitutive promoter for human elongation
factor EF-1-alpha"
misc_feature complement(6164..6280)
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 6311..6551
/label="U6 promoter"
/note="RNA polymerase III promoter for human U6 snRNA"
misc_RNA 6579..6654
/label="gRNA scaffold"
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
CDS complement(6743..6784)
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
CDS complement(6933..6977)
/label="gp41 peptide"
/note="antigenic peptide corresponding to amino acids 655
to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
al., 2013)"
misc_feature complement(7162..7395)
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
misc_feature complement(7888..8013)
/label="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
LTR 8060..8240
/label="5' LTR (truncated)"
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
promoter complement(8241..8467)
/label="RSV promoter"
/note="Rous sarcoma virus enhancer/promoter"