LOCUS       sequence-437460-        6835 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of human liver pyruvate kinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6835)
  AUTHORS   Swint-Kruse L, Dougherty LL, Page B, Wu T, O'Neil PT, Prasannan CB, 
            Timmons C, Tang Q, Parente DJ, Sreenivasan S, Holyoak T, Fenton AW
  TITLE     PYK-SubstitutionOME: an integrated database containing allosteric 
            coupling, ligand affinity and mutational, structural, pathological, 
            bioinformatic and computational information about pyruvate kinase 
            isozymes.
  JOURNAL   Database (Oxford). 2023 May 3;2023:baad030. doi: 
            10.1093/database/baad030.
  PUBMED    37171062
REFERENCE   2  (bases 1 to 6835)
  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: "Database 
            (Oxford)."; date: "2023-05-3"; volume: "2023:baad030. doi"; pages: "
            10.1093/database/baad030"
COMMENT     Sequence Label: pCDF-hLPYK
FEATURES             Location/Qualifiers
     source          1..6835
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        101..119
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    120..144
                     /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)."
     CDS             194..211
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1373..1393
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     promoter        3385..3403
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3404..3428
                     /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)."
     CDS             3537..3581
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(3619..3637)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3633..3680
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3851..4642)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(4643..4734)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4782..5520)
                     /direction=LEFT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     CDS             complement(5730..6812)
                     /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     6774..6793
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(join(6813..6835,1..55))
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga
       61 tctcgacgct ctcccttatg cgactcctgc attaggaaat taatacgact cactataggg
      121 gaattgtgag cggataacaa ttcccctgta gaaataattt tgtttaactt taataaggag
      181 atataccatg ggccatcatc atcatcacca cggtagcgcc ggcatgaaaa ttgaagaagg
      241 taaactggtt atttggatta atggcgataa aggctataat ggtctggccg aagttggtaa
      301 aaaatttgaa aaagataccg gtatcaaggt taccgttgaa catccggata aactggaaga
      361 aaaatttccg caggttgcag ccaccggtga cggccctgat attattttct gggcacatga
      421 tcgttttggc ggctatgccc agagcggcct gctggcagaa attaccccgg ataaagcctt
      481 tcaggataaa ctgtatccgt ttacctggga tgcagttcgt tataatggta aactgattgc
      541 ctatccgatt gcagtggaag cactgagcct gatctataat aaggatctgc tgccgaatcc
      601 gccgaaaacc tgggaagaaa ttccggcact ggataaagaa ctgaaagcca aaggtaaaag
      661 tgccctgatg tttaatctgc aggaaccgta ttttacctgg ccgctgattg cagcagatgg
      721 cggctatgcg tttaaatatg aaaatggcaa atacgatatt aaggacgtgg gtgttgataa
      781 tgccggtgcc aaagccggtc tgacctttct ggttgatctg attaagaata agcacatgaa
      841 tgcagatacc gattatagta ttgcagaagc cgcctttaat aagggcgaaa ccgccatgac
      901 cattaatggc ccgtgggcct ggagtaatat tgataccagc aaagtgaatt acggtgttac
      961 cgttctgccg acctttaaag gtcagccgag taaaccgttt gtgggtgtgc tgagcgcagg
     1021 cattaatgcc gccagtccga ataaggaact ggcaaaagaa tttctggaaa attatctgct
     1081 gaccgatgaa ggcctggaag cagtgaataa ggataaaccg ctgggtgcag tggccctgaa
     1141 aagctatgaa gaagaactgg ttaaagatcc gcgcattgca gcaacaatgg aaaatgccca
     1201 gaaaggcgaa attatgccga atattccgca gatgagcgca ttttggtatg ccgttcgtac
     1261 cgcagttatt aatgcagcca gcggccgtca gaccgttgat gaagccctga aagatgccca
     1321 gaccaatagt agtagtaata ataataacaa caacaacaat aacaacctgg gtgaaaatct
     1381 gtattttcag atggatagcg aagtgaatca ggaagcaaaa ccggaagtga aaccggaagt
     1441 taaaccggaa acccatatta atctgaaagt gagtgatggt agcagtgaaa ttttctttaa
     1501 aatcaagaag accaccccgc tgcgtcgcct gatggaagca tttgccaaac gtcagggtaa
     1561 agaaatggat agtctgcgtt ttctgtatga tggcattcgt attcaggccg atcaggcacc
     1621 ggaagatttg gatatggaag ataatgatat tatcgaggca catcgtgaac agattggtgg
     1681 catggaaggt ccggcaggtt atctgcgtcg tgccagtgtt gcccagctga cccaggaact
     1741 gggtacagca tttttccagc agcagcagct gccggcagca atggcagata cctttctgga
     1801 acatctgtgc ctgctggata ttgatagtga accggtggcc gcacgtagta ccagcattat
     1861 tgccaccatt ggtccggcca gtcgcagtgt ggaacgcctg aaagaaatga ttaaggcagg
     1921 tatgaatatc gcacgtctga attttagcca cggtagtcat gaatatcatg ccgaaagcat
     1981 tgccaatgtg cgtgaagccg tggaaagttt tgccggtagt ccgctgagct atcgcccggt
     2041 tgccattgca ctggatacca aaggtccgga aattcgcacc ggcattctgc agggcggtcc
     2101 ggaaagcgaa gttgaactgg ttaagggtag tcaggttctg gttaccgttg atccggcctt
     2161 tcgcacccgt ggtaatgcaa ataccgtttg ggttgattat ccgaatattg tgcgcgttgt
     2221 gccggtgggc ggtcgtatct atattgatga tggcctgatt agtctggttg tgcagaaaat
     2281 tggtccggaa ggcctggtta cccaggttga aaatggcggc gtgctgggta gtcgcaaagg
     2341 tgttaatctg ccgggtgccc aggttgatct gccgggtctg agtgaacagg atgttcgcga
     2401 tctgcgtttt ggtgtggaac atggcgtgga tattgtgttt gcaagttttg ttcgcaaagc
     2461 aagtgatgtg gccgccgttc gtgccgccct gggtcctgaa ggtcatggca ttaagattat
     2521 tagcaaaatt gagaaccacg aaggtgtgaa acgctttgat gaaattctgg aagttagcga
     2581 tggcattatg gttgcacgtg gtgacctggg cattgaaatt ccggccgaaa aagtttttct
     2641 ggcccagaaa atgatgattg gccgttgtaa tctggccggc aaaccggttg tgtgcgcaac
     2701 ccagatgctg gaaagcatga ttaccaaagc ccgcccgacc cgcgccgaaa ccagcgacgt
     2761 tgcaaatgcc gttctggatg gcgccgattg cattatgctg agcggcgaaa ccgcgaaagg
     2821 taattttccg gtggaagccg tgaaaatgca gcatgcaatt gcccgcgaag ccgaagccgc
     2881 cgtgtatcat cgtcagctgt ttgaagaact gcgccgcgcc gcaccgctga gtcgtgatcc
     2941 taccgaagtg accgcaattg gtgccgttga agcagcattc aaatgctgtg ccgcagcaat
     3001 tattgtgctg accaccaccg gtcgcagtgc acagctgctg agccgttatc gtccgcgtgc
     3061 cgccgttatt gcagtgaccc gtagcgcaca ggccgcccgt caggtgcatc tgtgtcgtgg
     3121 cgtttttccg ctgctgtatc gcgaaccgcc ggaagccatt tgggccgatg atgttgatcg
     3181 ccgtgtgcag tttggcattg aaagcggcaa actgcgtggc tttctgcgtg ttggcgatct
     3241 ggttattgtg gtgaccggct ggcgcccggg cagcggttat accaatatta tgcgtgtgct
     3301 gagcattagt taaaagcttg cggccgcata atgcttaagt cgaacagaaa gtaatcgtat
     3361 tgtacacggc cgcataatcg aaattaatac gactcactat aggggaattg tgagcggata
     3421 acaattcccc atcttagtat attagttaag tataagaagg agatatacat atggcagatc
     3481 tcaattggat atcggccggc cacgcgatcg ctgacgtcgg taccctcgag tctggtaaag
     3541 aaaccgctgc tgcgaaattt gaacgccagc acatggactc gtctactagc gcagcttaat
     3601 taacctaggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     3661 gggtcttgag gggttttttg ctgaaacctc aggcatttga gaagcacacg gtcacactgc
     3721 ttccggtagt caataaaccg gtaaaccagc aatagacata agcggctatt taacgaccct
     3781 gccctgaacc gacgaccggg tcatcgtggc cggatcttgc ggcccctcgg cttgaacgaa
     3841 ttgttagaca ttatttgccg actaccttgg tgatctcgcc tttcacgtag tggacaaatt
     3901 cttccaactg atctgcgcgc gaggccaagc gatcttcttc ttgtccaaga taagcctgtc
     3961 tagcttcaag tatgacgggc tgatactggg ccggcaggcg ctccattgcc cagtcggcag
     4021 cgacatcctt cggcgcgatt ttgccggtta ctgcgctgta ccaaatgcgg gacaacgtaa
     4081 gcactacatt tcgctcatcg ccagcccagt cgggcggcga gttccatagc gttaaggttt
     4141 catttagcgc ctcaaataga tcctgttcag gaaccggatc aaagagttcc tccgccgctg
     4201 gacctaccaa ggcaacgcta tgttctcttg cttttgtcag caagatagcc agatcaatgt
     4261 cgatcgtggc tggctcgaag atacctgcaa gaatgtcatt gcgctgccat tctccaaatt
     4321 gcagttcgcg cttagctgga taacgccacg gaatgatgtc gtcgtgcaca acaatggtga
     4381 cttctacagc gcggagaatc tcgctctctc caggggaagc cgaagtttcc aaaaggtcgt
     4441 tgatcaaagc tcgccgcgtt gtttcatcaa gccttacggt caccgtaacc agcaaatcaa
     4501 tatcactgtg tggcttcagg ccgccatcca ctgcggagcc gtacaaatgt acggccagca
     4561 acgtcggttc gagatggcgc tcgatgacgc caactacctc tgatagttga gtcgatactt
     4621 cggcgatcac cgcttccctc atactcttcc tttttcaata ttattgaagc atttatcagg
     4681 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagcta
     4741 gctcactcgg tcgctacgct ccgggcgtga gactgcggcg ggcgctgcgg acacatacaa
     4801 agttacccac agattccgtg gataagcagg ggactaacat gtgaggcaaa acagcagggc
     4861 cgcgccggtg gcgtttttcc ataggctccg ccctcctgcc agagttcaca taaacagacg
     4921 cttttccggt gcatctgtgg gagccgtgag gctcaaccat gaatctgaca gtacgggcga
     4981 aacccgacag gacttaaaga tccccaccgt ttccggcggg tcgctccctc ttgcgctctc
     5041 ctgttccgac cctgccgttt accggatacc tgttccgcct ttctccctta cgggaagtgt
     5101 ggcgctttct catagctcac acactggtat ctcggctcgg tgtaggtcgt tcgctccaag
     5161 ctgggctgta agcaagaact ccccgttcag cccgactgct gcgccttatc cggtaactgt
     5221 tcacttgagt ccaacccgga aaagcacggt aaaacgccac tggcagcagc cattggtaac
     5281 tgggagttcg cagaggattt gtttagctaa acacgcggtt gctcttgaag tgtgcgccaa
     5341 agtccggcta cactggaagg acagatttgg ttgctgtgct ctgcgaaagc cagttaccac
     5401 ggttaagcag ttccccaact gacttaacct tcgatcaaac cacctcccca ggtggttttt
     5461 tcgtttacag ggcaaaagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     5521 ttttctactg aaccgctcta gatttcagtg caatttatct cttcaaatgt agcacctgaa
     5581 gtcagcccca tacgatataa gttgtaattc tcatgttagt catgccccgc gcccaccgga
     5641 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     5701 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     5761 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5821 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     5881 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     5941 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     6001 actaccgaga tgtccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     6061 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     6121 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     6181 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     6241 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     6301 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca
     6361 gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc
     6421 tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc
     6481 accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca
     6541 cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc
     6601 agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg
     6661 cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca
     6721 gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac
     6781 tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg actct
//
