Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001380 | pmiRFP670-N1 | 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:
- pmiRFP670-N1
- Antibiotic Resistance:
- Kanamycin
- Length:
- 4960 bp
- Type:
- Mammalian Expression
- Replication origin:
- ori
- Selection Marker:
- Neomycin (select with G418)
- Copy Number:
- High Copy
- Promoter:
- CMV
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- CMV-F
- 3' Primer:
- SV40pA-R
pmiRFP670-N1 vector 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.
pmiRFP670-N1 vector Sequence
LOCUS 40924_30905 4960 bp DNA circular SYN 13-MAY-2021
DEFINITION Monomeric near-infrared fluorescent protein miRFP670 in mammalian
plasmid.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4960)
AUTHORS Shcherbakova DM, Baloban M, Emelyanov AV, Brenowitz M, Guo P,
Verkhusha VV
TITLE Bright monomeric near-infrared fluorescent proteins as tags and
biosensors for multiscale imaging.
JOURNAL Nat Commun. 2016 Aug 19;7:12405. doi: 10.1038/ncomms12405.
PUBMED 27539380
REFERENCE 2 (bases 1 to 4960)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 4960)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nat
Commun."; date: "2016-08-19"; pages: "
10.1038/ncomms12405"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..4960
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 106..409
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 410..613
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
misc_feature 636..716
/label=MCS
/note="multiple cloning site"
CDS 724..1668
/codon_start=1
/label=ORF frame 1
/note="bright monomeric near-infrared fluorescent protein
engineered from a bacterial phytochrome, with an emission
maximum of 670 nm (Shcherbakova et al., 2016)"
/translation="MVAGHASGSPAFGTASHSNCEHEEIHLAGSIQPHGALLVVSEHDH
RVIQASANAAEFLNLGSVLGVPLAEIDGDLLIKILPHLDPTAEGMPVAVRCRIGNPSTE
YCGLMHRPPEGGLIIELERAGPSIDLSGTLAPALERIRTAGSLRALCDDTVLLFQQCTG
YDRVMVYRFDEQGHGLVFSECHVPGLESYFGNRYPSSTVPQMARQLYVRQRVRVLVDVT
YQPVPLEPRLSPLTGRDLDMSGCFLRSMSPCHLQFLKDMGVRATLAVSLVVGGKLWGLV
VCHHYLPRFIRFELRAICKRLAERIATRITALES"
polyA_signal 1793..1914
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin complement(1921..2376)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 2403..2507
/label=AmpR promoter
promoter 2509..2866
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
CDS 2901..3692
/codon_start=1
/label=NeoR/KanR
/note="aminoglycoside phosphotransferase"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(3883..3902)
/label=TK-pA-R
/note="Thymidine kinase polyA, reverse primer"
polyA_signal 3927..3974
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
rep_origin 4303..4891
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"