Human TAS2R4 ORF clone (NM_016944) (Cat. No.: V044221)
- Name:
- pCDNA5-FRT-To-TAS2R4(human)-HSV tag
- Accession ID:
- NM_016944.2
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6131 bp
- Type:
- Protein expression
- Replication origin:
- ori
- Host:
- Mammalian cells
- Selection Marker:
- Hyg
- Copy Number:
- High
- Promoter:
- CMV/to
- 5' Primer:
- CMV-F
- Fusion Tag:
- HSV tag
- Growth Strain(s):
- DH5a
- Growth Temperature:
- 37℃
- Expression Method:
- Transient
- Gene Synonyms:
- T2R4
- Transcript Definition:
- Homo sapiens taste 2 receptor member 4 (TAS2R4), mRNA
Resources
Plasmid Protocol
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add sterile water to dissolve the DNA: add 20 μl for 5 μg plasmid, and 100 μl for 100 μg plasmid.
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.
Human TAS2R4 ORF clone (NM_016944) (Cat. No.: V044221) Sequence
LOCUS V044221 6131 bp DNA circular SYN 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION V044221
VERSION V044221
KEYWORDS .
SOURCE .
ORGANISM .
.
FEATURES Location/Qualifiers
enhancer 245..624
/note="human cytomegalovirus immediate early enhancer"
/label="CMV enhancer"
promoter 625..828
/note="human cytomegalovirus (CMV) immediate early
promoter"
/label="CMV promoter"
promoter 873..891
/note="promoter for bacteriophage T7 RNA polymerase"
/label="T7 promoter"
regulatory 911..920
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
/label="Kozak sequence"
CDS 1058..1954
/label="TAS2R4(NM_016944)"
/note="TAS2R4(NM_016944)"
/gene="TAS2R4"
CDS 1961..1993
/codon_start=1
/product="HSV (herpes simplex virus) epitope tag"
/transl_table=1
/translation="QPELAPEDPED"
/label="HSV tag"
polyA_signal 2099..2323
/note="bovine growth hormone polyadenylation signal"
/label="bGH poly(A) signal"
protein_bind 2607..2654
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2μ plasmid"
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
/label="FRT"
CDS 2662..3684
/codon_start=1
/gene="aph(4)-Ia"
/note="confers resistance to hygromycin"
/product="aminoglycoside phosphotransferase from E. coli"
/transl_table=1
/translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
*"
/label="HygR"
polyA_signal 3814..3935
/note="SV40 polyadenylation signal"
/label="SV40 poly(A) signal"
primer_bind complement(3984..4000)
/note="common sequencing primer, one of multiple similar
variants"
/label="M13 rev"
protein_bind 4008..4024
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-β-D-thiogalactopyranoside (IPTG)."
/label="lac operator"
promoter complement(4032..4062)
/note="promoter for the E. coli lac operon"
/label="lac promoter"
protein_bind 4077..4098
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
/label="CAP binding site"
rep_origin complement(4386..4974)
/direction=LEFT
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
/label="ori"
CDS complement(5145..6005)
/codon_start=1
/gene="bla"
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/product="β-lactamase"
/transl_table=1
/translation="MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGY
IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY
SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR
WEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS
ALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA
SLIKHW*"
/label="AmpR"
promoter complement(6006..6110)
/gene="bla"
/label="AmpR promoter"