pCold- GST Plasmid

$216.00

  • Model: PVT0608
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pCold-GST

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pCold-GST,Plasmid pCold-GST,pCold-GST vector

 

pCold-GST Information

Promoter: CSPA

Replicon: ColE1 ori

Plasmid classification: Escherichia coli vector; pCold series expression plasmid

Plasmid size: 5097bp

Plasmid labeling: N-GST, N-HRV 3C protease cutting site, N-His

Prokaryotic resistance: ampicillin Amp

Cloned strain: Escherichia coli DH5 alpha

Culture conditions: 37 centigrade, aerobic, LB

Expression host: Escherichia coli

Culture conditions: 37 centigrade, aerobic, LB

Inducement: IPTG or lactose and its analogues

5'sequencing primers: pCold Forward: 5-ACGCCATATCGCCGAAAGG-3

3'sequencing primers: pCold Reverse 5-GGCAGGGATCTTAGATTCTG-3

Note: the pCold-GST vector expresses GST fusion protein, and the GST label can improve the soluble protein of the target protein and improve the purity of the target protein.

 

pCold-GST Description

Based on the cold shock vector, pCold-GST DNA integrated glutathione S- transferase (glutathione S-transferases, GST) soluble label from Schistosoma japonicum. The fusion expression of GST at the end of the target protein N can improve the stability and solubility of the fusion protein. This product inserts 5 'non coding region (5' -UTR), translation enhancer element (TEE), His tag, GST tag and polyclonal site (MCS) at the downstream of cspA promoter. In addition, the lac operator, which can strictly regulate the expression of the target gene, is also inserted at the lower reaches of the cspA promoter. GST tag fusion protein can be purified for high affinity. The recognition sequence of high specific HRV 3C Protease was inserted between the GST label and the polyclonal site (MCS), which could remove the label from the fusion protein. The optimum temperature of HRV 3C Protease is 4~5 C, and the label removal reaction of the target protein can be carried out under mild conditions. The promoter of the pCold series is derived from Escherichia coli, so most Escherichia coli can be used as an expression host.

pCold-GST-polyclonal-site

 

pCold-GST Sequence

LOCUS       Exported                5097 bp ds-DNA     circular SYN 06-JUL-2017
DEFINITION  synthetic circular DNA
FEATURES             Location/Qualifiers
     source          1..5097
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        86..163
                     /gene="lacI"
                     /label=lacI promoter
     CDS             164..1246
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /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-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     rep_origin      complement(1418..2006)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2177..3037)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(3038..3142)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3242..3697
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     CDS             complement(4111..4134)
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(4141..4794)
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             complement(4798..4809)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(4810..4827)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    4998..5014
                     /label=lac operator
                     /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-beta-D-thiogalactopyranoside (IPTG)."
ORIGIN
        1 gcggcggcgg tgctcaacgg cctcaaccta ctactgggct gcttcctaat gcaggagtcg
       61 cataagggag agcgtcgaga tcccggacac catcgaatgg cgcaaaacct ttcgcggtat
      121 ggcatgatag cgcccggaag agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt
      181 atacgatgtc gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca
      241 ggccagccac gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa
      301 ttacattccc aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt
      361 tgccacctcc agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg
      421 cgccgatcaa ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc
      481 ctgtaaagcg gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga tcattaacta
      541 tccgctggat gaccaggatg ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt
      601 atttcttgat gtctctgacc agacacccat caacagtatt attttctccc atgaagacgg
      661 tacgcgactg ggcgtggagc atctggtcgc attgggtcac cagcaaatcg cgctgttagc
      721 gggcccatta agttctgtct cggcgcgtct gcgtctggct ggctggcata aatatctcac
      781 tcgcaatcaa attcagccga tagcggaacg ggaaggcgac tggagtgcca tgtccggttt
      841 tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc actgcgatgc tggttgccaa
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