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pSFV-Helper1 vector (V000444)

Price Information

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V000444 pSFV-Helper1 In stock, instant shipping

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

Basic Vector Information

pSFV-Helper1 is a plasmid for transcription of defective helper RNA, providing structural Semliki forest virus (SFV) proteins to package recombinant SFV genomes into virus like particles.

Vector Name:
pSFV-Helper1
Antibiotic Resistance:
Ampicillin
Length:
8192 bp
Type:
Helper plasmid, for run-off transcription and RNA
Replication origin:
ori
Promoter:
SP6
Growth Strain(s):
DH5alpha
Growth Temperature:
37℃

pSFV-Helper1 vector Map

Copy Sequence View Map
pSFV-Helper18192 bp400800120016002000240028003200360040004400480052005600600064006800720076008000SP6 promoterStructural polyproteinL4440oriAmpRAmpR promoterpBRforEcopGEX 3'pRS-markerpBRrevBam

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

  • Liljeström P, Garoff H. A new generation of animal cell expression vectors based on the Semliki Forest virus replicon. Biotechnology (N Y). 1991 Dec;9(12):1356-61.

pSFV-Helper1 vector Sequence

LOCUS       V000444                 8192 bp    DNA     circular SYN 13-MAY-2021
DEFINITION  Exported.
ACCESSION   V000444
VERSION     V000444
KEYWORDS    pSFV-Helper1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8192)
  AUTHORS   Liljestrom P, Garoff H
  TITLE     A new generation of animal cell expression vectors based on the
            Semliki Forest virus replicon.
  JOURNAL   Biotechnology (N Y). 1991 Dec;9(12):1356-61.
   PUBMED   1370252
REFERENCE   2  (bases 1 to 8192)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 8192)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName:
            "Biotechnology (N Y)."; date: "1991-12"; volume: "9(12)"; pages:
            "1356-61"
            SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8192
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..19
                     /label="SP6 promoter"
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     CDS             1347..5105
                     /note="Structural polyprotein from Semliki forest virus.
                     Accession#: P03315"
                     /label="Structural polyprotein"
     primer_bind     complement(5510..5527)
                     /label="L4440"
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5681..6269)
                     /direction=LEFT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(6443..7300)
                     /label="AmpR"
                     /note="beta-lactamase"
     promoter        complement(7301..7405)
                     /label="AmpR promoter"
     primer_bind     7473..7491
                     /label="pBRforEco"
                     /note="pBR322 vectors, upsteam of EcoRI site, forward
                     primer"
     primer_bind     complement(7529..7551)
                     /label="pGEX 3'"
                     /note="pGEX vectors, reverse primer"
     primer_bind     7651..7670
                     /label="pRS-marker"
                     /note="pRS vectors, use to sequence yeast selectable
                     marker"
     primer_bind     7884..7903
                     /label="pBRrevBam"
                     /note="pBR322 vectors, tet region, downstream of BamHI,
                     reverse primer"