Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V017588 | pEGFP-VSVG | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
pEGFP-VSVG enabled the visualisation of membrane transport from the endoplasmic reticulum to the Golgi apparatus in living cells via fluorescence technology, revealing the transport mechanism of non-vesicular precursor structures and their dependence on microtubules and dynein/dynein-activating factors.
- Vector Name:
- pEGFP-VSVG
- Antibiotic Resistance:
- Kanamycin
- Length:
- 6254 bp
- Type:
- Protein expression
- Replication origin:
- ori
- Host:
- Mammalian cells
- Selection Marker:
- Neo/G418
- Promoter:
- CMV
- 5' Primer:
- CMV-F
- Growth Strain(s):
- JM109
- Growth Temperature:
- 37℃
pEGFP-VSVG 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.
References
- Rauter T, Burgstaller S, Gottschalk B, Ramadani-Muja J, Bischof H, Hay JC, Graier WF, Malli R. ER-to-Golgi Transport in HeLa Cells Displays High Resilience to Ca2+ and Energy Stresses. Cells. 2020 Oct 17;9(10):2311. doi: 10.3390/cells9102311. PMID: 33080790; PMCID: PMC7603030.
pEGFP-VSVG vector Sequence
LOCUS . 6254 bp DNA circular UNK 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION
VERSION
KEYWORDS .
SOURCE .
ORGANISM .
.
FEATURES Location/Qualifiers
enhancer 61..364
/label="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
promoter 365..568
/label="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
CDS 625..2157
/label="VSV-G tsO45"
/note="vesicular stomatitis virus G glycoprotein,
thermosensitive tsO45 mutant"
CDS 2200..2916
/label="mEGFP"
/note="enhanced GFP with monomerizing A206K mutation
(Zacharias et al., 2002)"
polyA_signal 3042..3163
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
rep_origin complement(3170..3625)
/label="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 3652..3756
/label="AmpR promoter"
promoter 3758..4115
/label="SV40 promoter"
/note="SV40 enhancer and early promoter"
CDS 4150..4941
/label="NeoR/KanR"
/note="aminoglycoside phosphotransferase from Tn5"
polyA_signal 5176..5223
/label="HSV TK poly(A) signal"
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
rep_origin 5552..6140
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"