Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V012492 | pFN19K HaloTag T7 SP6 | In stock, instant shipping |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
The pFN19K HaloTag T7 SP6 vector is equipped with a kanamycin resistance gene and is designed for cell-free expression. It enables the production of a protein featuring an N-terminal HaloTag that can be cleaved.
- Vector Name:
- pFN19K HaloTag T7 SP6
- Antibiotic Resistance:
- Kanamycin
- Length:
- 4421 bp
- Type:
- Cloning Vectors
- Replication origin:
- ori
- Source/Author:
- Promega
- Selection Marker:
- Neomycin/G418(Geneticin)
- Copy Number:
- High copy number
- Promoter:
- SP6
- Growth Strain(s):
- Top10
- Growth Temperature:
- 37℃
pFN19K HaloTag T7 SP6 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.
pFN19K HaloTag T7 SP6 vector Sequence
LOCUS 40924_20301 4421 bp DNA circular SYN 01-JAN-1980
DEFINITION Flexi(R) vector with a kanamycin resistance marker, for cell-free
expression of a protein with a cleavable N-terminal HaloTag(R).
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4421)
AUTHORS Promega
TITLE Direct Submission
REFERENCE 2 (bases 1 to 4421)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
COMMENT Subclone a coding sequence between the SgfI/AsiSI and PmeI sites to
remove the lethal barnase gene.
FEATURES Location/Qualifiers
source 1..4421
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 21..39
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 45..63
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
CDS 80..970
/codon_start=1
/label=HaloTag(R)
/note="modified bacterial dehalogenase that forms covalent
bonds with chloroalkane derivatives"
/translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
IGSEIARWLSTLEISG"
CDS 983..1003
/codon_start=1
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
/translation="EDLYFQS"
CDS 1041..1373
/codon_start=1
/label=barnase
/note="ribonuclease from Bacillus amyloliquefaciens"
/translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
QTFTKIR"
misc_feature 1386..1441
/label=MCS
/note="pUC18/19 multiple cloning site"
terminator 1544..1591
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
CDS 1972..2763
/codon_start=1
/label=NeoR/KanR
/note="aminoglycoside phosphotransferase from Tn5"
/translation="MLEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
rep_origin complement(2938..3526)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature 3642..3925
/label=cer region
/note="ColE1-derived recombination site that helps to
maintain plasmids as monomers"
terminator 4155..4241
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 4333..4360
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"