Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V018120 | pN1-cAMPinG1 | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
Expression vector of genetically encoded cAMP indicator, cAMPinG1. cAMPinG1 under the control of a CAG promoter with expression enhanced by WPRE (woodchuck hepatitis virus posttranscriptional regulatory element).
- Vector Name:
- pN1-cAMPinG1
- Antibiotic Resistance:
- Kanamycin
- Length:
- 7368 bp
- Type:
- BLE84607
- Source/Author:
- Tatsushi Yokoyama
- Growth Strain(s):
- Top10
- Growth Temperature:
- 37℃
pN1-cAMPinG1 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
- Yokoyama T, Manita S, Uwamori H, Tajiri M, Imayoshi I, Yagishita S, Murayama M, Kitamura K, Sakamoto M. A multicolor suite for deciphering population coding of calcium and cAMP in vivo. Nat Methods. 2024 May;21(5):897-907. doi: 10.1038/s41592-024-02222-9. Epub 2024 Mar 21. PMID: 38514778; PMCID: PMC11093745.
pN1-cAMPinG1 vector Sequence
LOCUS Exported 7368 bp DNA circular SYN 22-MAR-2025
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7368)
AUTHORS RIKEN DNA Bank
TITLE Direct Submission
COMMENT RDB19691_pN1-cAMPinG1
FEATURES Location/Qualifiers
source 1..7368
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 95..1694
/label=CAG promoter
enhancer 95..398
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 400..677
/label=chicken beta-actin promoter
intron 678..1694
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
CDS 1803..3458
/codon_start=1
/label=cAMPinG1
/label=CyA033
/translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDEDAASYVR
KVIPKDYKTMAALAKAIEKNVLFSHLDDNERSDIFDAMFPVSFIAGETVIQQGDEGDNF
YVIDQGEMDVYVSWPVYIKADKQKNGIKANFKIWHNIEDGGVQLAYHYQQNTPIGDGPV
LLPDNHYLLVQSSLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEEL
FTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTY
GVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIEL
KGIDFKEDGNILGHKLEYNGWATSVGEGGSFGELALIYGTPRAATVKAKTNVKLWGIDR
DSYRRILMGSTLRKRKMYEEFLSKVSILESLDKWERLTVADALEPVQFEDGQKIVVQGE
PGDEFFIILEGTAAVLQRRSENEEFVEVGRLGPSDYFGEIALLMNRPRAATVVARGPLK
CVKLDRPRFERVLGPCSDILKRNIQQYNSFVSLSV"
CDS 1812..1829
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 1833..1865
/codon_start=1
/product="leader peptide from bacteriophage T7 gene 10"
/label=T7 tag (gene 10 leader)
/note="promotes efficient translation in E. coli"
/translation="MASMTGGQQMG"
CDS 1869..1892
/codon_start=1
/product="Xpress(TM) epitope tag, including an enterokinase
recognition and cleavage site"
/label=Xpress(TM) tag
/translation="DLYDDDDK"
misc_feature 3493..4081
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
polyA_signal 4110..4231
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin complement(4331..4786)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 4813..4917
/gene="bla"
/label=AmpR promoter
promoter 4919..5276
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
enhancer 4919..5110
/label=SV40 enhancer
/note="enhancer for the SV40 early promoter (Herr, 1993)"
rep_origin 5127..5262
/label=SV40 ori
/note="SV40 origin of replication"
CDS 5311..6105
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
polyA_signal 6337..6384
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
misc_feature 6713..7301
/label=pUC ori