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pFA6a-5FLAG-hphMX6 vector (V006371)

Price Information

Cat No. Plasmid Name Availability Add to cart
V006371 pFA6a-5FLAG-hphMX6 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:
pFA6a-5FLAG-hphMX6
Antibiotic Resistance:
Ampicillin
Length:
4531 bp
Type:
Tagging vector
Replication origin:
ori
Host:
Yeast
Source/Author:
Noguchi C, Garabedian MV, Malik M, Noguchi E.
Promoter:
TEF

pFA6a-5FLAG-hphMX6 vector Vector Map

pFA6a-5FLAG-hphMX64531 bp6001200180024003000360042005xFLAGADH1 terminatorhphMX6T7 promoteroriAmpRAmpR promoterSP6 promoter

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.

pFA6a-5FLAG-hphMX6 vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       40924_19211        4531 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Tagging vector pFA6a-5FLAG-hphMX6, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4531)
  AUTHORS   Noguchi C, Garabedian MV, Malik M, Noguchi E.
  TITLE     A vector system for genomic FLAG epitope-tagging in 
            Schizosaccharomyces pombe
  JOURNAL   Biotechnol J 3 (9-10), 1280-1285 (2008)
  PUBMED    18729046
REFERENCE   2  (bases 1 to 4531)
  AUTHORS   Noguchi C, Garabedian M, Malik M, Noguchi E.
  TITLE     Direct Submission
  JOURNAL   Submitted (16-FEB-2008) Biochemistry and Molecular Biology, Drexel 
            University College of Medicine, 245 N 15th Street, MS 497, 
            Philadelphia, PA 19102, USA
REFERENCE   3  (bases 1 to 4531)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 4531)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Biotechnol 
            J 3 (9-10), 1280-1285 (2008)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (16-FEB-2008) Biochemistry and Molecular Biology, Drexel University 
            College of Medicine, 245 N 15th Street, MS 497, Philadelphia, PA 
            19102, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4531
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             69..200
                     /codon_start=1
                     /label=5xFLAG
                     /note="five tandem FLAG(R) epitope tags"
                     /translation="DYKDDDDKDYKDDDDKLMDYKDDDDKDYKDDDDKLMDYKDDDDK"
     terminator      227..414
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae
                     alcohol dehydrogenase 1 (ADH1) gene"
     gene            489..2064
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin
                     resistance"
     promoter        complement(2169..2187)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      complement(2445..3033)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3207..4064)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(4065..4169)
                     /label=AmpR promoter
     promoter        4515..4531
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"