Human ALK ORF clone (NM_004304) (Cat. No.: V043865)

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Basic Information

Name:: pCDNA5-FRT-To-ALK(human)

Accession ID:: NM_004304.5

Antibiotic Resistance:: Ampicillin

Length:: 9989 bp

Type:: Protein expression

Replication origin:: ori

Host:: Mammalian cells

Selection Marker:: Hyg

Copy Number:: High

Promoter:: CMV/To

5' Primer:: CMV-F

Growth Strain(s):: DH5a

Growth Temperature:: 37℃

Expression Method:: Transient

Gene Synonyms:: ALK1; CD246; NBLST3

Transcript Definition:: Homo sapiens ALK receptor tyrosine kinase (ALK), transcript variant 1, mRNA

Cat. No.: V043865 pCDNA5-FRT-To-ALK(human)

$ 299.0

In stock, 1-2 weeks for quality controls

Two tubes of transfection grade plasmid, each tube is about 100 µg (lyophilized).

Transcript ID

NM_004304.5

Fusion Tag

3xFLAG

Selection Marker

Hyg Neo/G418 Puro;CopGFP

Expression Method

Transient Constiutive, Stable / Transient

Resources

Plasmid Protocol

Plasmid Reconstitution Protocol

Centrifuge at 5,000×g for 5 min.
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.
Close the tube and incubate for 10 minutes at room temperature.
Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
Store the plasmid at -20 ℃.
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

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.
Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
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.
Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
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 ALK ORF clone (NM_004304) (Cat. No.: V043865) Annotation

LOCUS       V043865                 9989 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V043865
VERSION     V043865
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        235..614
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        615..818
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     protein_bind    820..838
                     /bound_moiety="tetracycline repressor TetR"
                     /gene="tetO"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /label="tet operator"
     protein_bind    841..859
                     /bound_moiety="tetracycline repressor TetR"
                     /gene="tetO"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /label="tet operator"
     CDS             1008..5867
                     /label="ALK(NM_004304)"
                     /note="ALK(NM_004304)"
                     /gene="ALK"
     polyA_signal    5947..6171
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     protein_bind    6455..6502
                     /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             6510..7532
                     /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    7662..7783
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(7832..7848)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    7856..7872
                     /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(7880..7910)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    7925..7946
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(8234..8822)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8993..9853)
                     /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(9854..9958)
                     /gene="bla"
                     /label="AmpR promoter"