Human TM6SF2 ORF clone (NM_001001524) (Cat. No.: V021769)
- Name:
- pLVX-TetOne-Puro-TM6SF2(human)
- Accession ID:
- NM_001001524.3
- Antibiotic Resistance:
- Ampicillin
- Length:
- 10363 bp
- Type:
- Protein expression, Tetracycline inducible
- Replication origin:
- ori
- Host:
- Mammalian cells, Lentivirus
- Selection Marker:
- Puro
- Promoter:
- TRE3GS
- Expression Method:
- Tetracycline inducible
- Transcript Definition:
- Homo sapiens transmembrane 6 superfamily member 2 (TM6SF2), mRNA
Resources
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.
Human TM6SF2 ORF clone (NM_001001524) (Cat. No.: V021769) Sequence
LOCUS V021769 10363 bp DNA circular SYN 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION V021769
VERSION V021769
KEYWORDS .
SOURCE .
ORGANISM .
.
FEATURES Location/Qualifiers
polyA_signal 216..350
/note="SV40 polyadenylation signal"
/label="SV40 poly(A) signal"
promoter 398..502
/gene="bla"
/label="AmpR promoter"
CDS 503..1363
/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"
rep_origin 1534..2119
/direction=RIGHT
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
/label="ori"
protein_bind 2407..2428
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
/label="CAP binding site"
promoter 2443..2473
/note="promoter for the E. coli lac operon"
/label="lac promoter"
protein_bind 2481..2497
/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"
primer_bind 2505..2521
/note="common sequencing primer, one of multiple similar
variants"
/label="M13 rev"
LTR 2650..3283
/note="3' long terminal repeat (LTR) from HIV-1"
/label="3' LTR"
misc_feature 3491..4079
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
/label="WPRE"
CDS complement(4093..4692)
/codon_start=1
/gene="pac from Streptomyces alboniger"
/note="confers resistance to puromycin"
/product="puromycin N-acetyltransferase"
/transl_table=1
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA*"
/label="PuroR"
promoter complement(4701..5030)
/note="SV40 enhancer and early promoter"
/label="SV40 promoter"
CDS complement(5041..5787)
/codon_start=1
/product="modified rtTA protein that binds tightly to
promoters containing the tet operator in the presence of
doxycycline"
/transl_table=1
/translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
DALDDFDLDMLPADALDDFDLDMLPG*"
/label="Tet-On® 3G"
promoter complement(5806..6316)
/note="human phosphoglycerate kinase 1 promoter"
/label="hPGK promoter"
promoter 6333..6700
/note="3rd-generation Tet-responsive promoter that can be
activated by binding of Tet-On® 3G, modified to eliminate
binding sites for endogenous mammalian transcription
factors"
/label="TRE3GS promoter"
CDS 6729..7859
/label="TM6SF2(NM_001001524)"
/note="TM6SF2(NM_001001524)"
/gene="TM6SF2"
polyA_signal 8043..8177
/note="SV40 polyadenylation signal"
/label="SV40 poly(A) signal"
misc_feature 8221..8336
/note="central polypurine tract and central termination
sequence of HIV-1 (lacking the first T)"
/label="cPPT/CTS"
CDS complement(8599..8643)
/codon_start=1
/note="recognized by the 2H10 single-chain llama nanobody"
/product="antigenic peptide corresponding to amino acids
655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
et al., 2013)"
/transl_table=1
/translation="KNEQELLELDKWASL"
/label="gp41 peptide"
misc_feature 8828..9061
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
/label="RRE"
misc_feature 9558..9683
/note="packaging signal of human immunodeficiency virus
type 1"
/label="HIV-1 Ψ"
LTR 9730..10363
/note="3' long terminal repeat (LTR) from HIV-1"
/label="3' LTR"