LOCUS       sequence-238811-        6925 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express His-tagged PKM2 (wild type) protein in bacteria..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6925)
  AUTHORS   Chen TJ, Wang HJ, Liu JS, Cheng HH, Hsu SC, Wu MC, Lu CH, Wu YF, Wu 
            JW, Liu YY, Kung HJ, Wang WC
  TITLE     Mutations in the PKM2 exon-10 region are associated with reduced 
            allostery and increased nuclear translocation.
  JOURNAL   Commun Biol. 2019 Mar 15;2:105. doi: 10.1038/s42003-019-0343-4. 
            eCollection 2019.
  PUBMED    30911680
REFERENCE   2  (bases 1 to 6925)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Commun 
            Biol."; date: "2019-03-15"; volume: "2:105. doi"; pages: " 
            10.1038/s42003-019-0343-4. eCollection 2019"
COMMENT     Sequence Label: pET28a-PKM2-WT
FEATURES             Location/Qualifiers
     source          1..6925
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..18
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1664..1681
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1734..1752)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1748..1795
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1832..2287
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1919..1938)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2129..2150
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2379..3194)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2379..3194)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3102..3121
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3186..3205
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3316..3904
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3805..3824
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4058..4075
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4090..4229
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4155..4174)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4315..4337
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4331..4522)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5331..6413)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     6375..6394
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6414..6491)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6697..6716
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6800..6819
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6800..6818
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6819..6843
                     /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)."
     RBS             6858..6880
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             6899..6916
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 ctggtgccgc gcggcagcca tatgtcgaag ccccatagtg aagccgggac tgccttcatt
       61 cagacccagc agctgcacgc agccatggct gacacattcc tggagcacat gtgccgcctg
      121 gacattgatt caccacccat cacagcccgg aacactggca tcatctgtac cattggccca
      181 gcttcccgat cagtggagac gttgaaggag atgattaagt ctggaatgaa tgtggctcgt
      241 ctgaacttct ctcatggaac tcatgagtac catgcggaga ccatcaagaa tgtgcgcaca
      301 gccacggaaa gctttgcttc tgaccccatc ctctaccggc ccgttgctgt ggctctagac
      361 actaaaggac ctgagatccg aactgggctc atcaagggca gcggcactgc agaggtggag
      421 ctgaagaagg gagccactct caaaatcacg ctggataacg cctacatgga aaagtgtgac
      481 gagaacatcc tgtggctgga ctacaagaac atctgcaagg tggtggaagt gggcagcaag
      541 atctacgtgg atgatgggct tatttctctc caggtgaagc agaaaggtgc cgacttcctg
      601 gtgacggagg tggaaaatgg tggctccttg ggcagcaaga agggtgtgaa ccttcctggg
      661 gctgctgtgg acttgcctgc tgtgtcggag aaggacatcc aggatctgaa gtttggggtc
      721 gagcaggatg ttgatatggt gtttgcgtca ttcatccgca aggcatctga tgtccatgaa
      781 gttaggaagg tcctgggaga gaagggaaag aacatcaaga ttatcagcaa aatcgagaat
      841 catgaggggg ttcggaggtt tgatgaaatc ctggaggcca gtgatgggat catggtggct
      901 cgtggtgatc taggcattga gattcctgca gagaaggtct tccttgctca gaagatgatg
      961 attggacggt gcaaccgagc tgggaagcct gtcatctgtg ctactcagat gctggagagc
     1021 atgatcaaga agccccgccc cactcgggct gaaggcagtg atgtggccaa tgcagtcctg
     1081 gatggagccg actgcatcat gctgtctgga gaaacagcca aaggggacta tcctctggag
     1141 gctgtgcgca tgcagcacct gattgcccgt gaggcagagg ctgccatcta ccacttgcaa
     1201 ttatttgagg aactccgccg cctggcgccc attaccagcg accccacaga agccaccgcc
     1261 gtgggtgccg tggaggcctc cttcaagtgc tgcagtgggg ccataatcgt cctcaccaag
     1321 tctggcaggt ctgctcacca ggtggccaga taccgcccac gtgcccccat cattgctgtg
     1381 acccggaatc cccagacagc tcgtcaggcc cacctgtacc gtggcatctt ccctgtgctg
     1441 tgcaaggacc cagtccagga ggcctgggct gaggacgtgg acctccgggt gaactttgcc
     1501 atgaatgttg gcaaggcccg aggcttcttc aagaagggag atgtggtcat tgtgctgacc
     1561 ggatggcgcc ctggctccgg cttcaccaac accatgcgtg ttgttcctgt gccgtgagga
     1621 tccgaattcg agctccgtcg acaagcttgc ggccgcactc gagcaccacc accaccacca
     1681 ctgagatccg gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga
     1741 gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa
     1801 aggaggaact atatccggat tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg
     1861 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     1921 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     1981 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     2041 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     2101 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     2161 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     2221 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt
     2281 acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     2341 aaatacattc aaatatgtat ccgctcatga attaattctt agaaaaactc atcgagcatc
     2401 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
     2461 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
     2521 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
     2581 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
     2641 agtttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
     2701 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg
     2761 cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
     2821 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
     2881 gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
     2941 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     3001 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     3061 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     3121 ccatataaat cagcatccat gttggaattt aatcgcggcc tagagcaaga cgtttcccgt
     3181 tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt
     3241 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     3301 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     3361 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     3421 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     3481 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     3541 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     3601 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     3661 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     3721 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     3781 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     3841 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     3901 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     3961 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     4021 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     4081 ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcaa
     4141 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc
     4201 tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc
     4261 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     4321 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa
     4381 gctcatcagc gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct
     4441 cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg
     4501 cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg
     4561 taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg
     4621 cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga
     4681 gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg
     4741 gtagccagca gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc
     4801 gcgtttccag actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg
     4861 cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct
     4921 aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc
     4981 gcacccgtgg ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg
     5041 cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca
     5101 ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg
     5161 ccggcacctg tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag
     5221 tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc
     5281 gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg
     5341 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     5401 gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac
     5461 agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt
     5521 tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg
     5581 tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg
     5641 gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga
     5701 acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg
     5761 ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc
     5821 agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga
     5881 cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata
     5941 ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca
     6001 gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg
     6061 cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc
     6121 atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt
     6181 tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg
     6241 cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc
     6301 actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc
     6361 tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc
     6421 accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat
     6481 tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc
     6541 cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat
     6601 ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct
     6661 catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc
     6721 agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga
     6781 gatctcgatc ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat
     6841 tcccctctag aaataatttt gtttaacttt aagaaggaga tataccatgg gcagcagcca
     6901 tcatcatcat catcacagca gcggc
//
