LOCUS       sequence-76071-e        4340 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  overexpression of the human TPI1 Ile170Val allele in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4340)
  AUTHORS   Gruning NM, Du D, Keller MA, Luisi BF, Ralser M
  TITLE     Inhibition of triosephosphate isomerase by phosphoenolpyruvate in 
            the feedback-regulation of glycolysis.
  JOURNAL   Open Biol. 2014 Mar 5;4(3):130232. doi: 10.1098/rsob.130232.
  PUBMED    24598263
REFERENCE   2  (bases 1 to 4340)
  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: "Open 
            Biol."; date: "2014-03-5"; volume: "4(3):130232. doi"; pages: " 
            10.1098/rsob.130232"
COMMENT     Sequence Label: pET20b- hTPI Ile170Val
FEATURES             Location/Qualifiers
     source          1..4340
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     342..361
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        342..360
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             392..414
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1178..1195
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1248..1266)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1262..1309
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1346..1801
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1433..1452)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1643..1664
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1828..1932
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1933..2001,2002..2793)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2151..2170)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2964..3552
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3453..3472
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3706..3723
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3738..3880
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3806..3825)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3966..3988
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3982..4173)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc
       61 agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca
      121 gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt
      181 tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa
      241 ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg
      301 gccaggaccc aacgctgccc gagatctcga tcccgcgaaa ttaatacgac tcactatagg
      361 gagaccacaa cggtttccct ctagaaataa ttttgtttaa ctttaagaag gagatataca
      421 tatgatggcg ccctccagga agttcttcgt tgggggaaac tggaagatga acgggcggaa
      481 gcagagtctg ggggagctca tcggcactct gaacgcggcc aaggtgccgg ccgacaccga
      541 ggtggtttgt gctcccccta ctgcctatat cgacttcgcc cggcagaagc tagatcccaa
      601 gattgctgtg gctgcgcaga actgctacaa agtgactaat ggggctttta ctggggagat
      661 cagccctggc atgatcaaag actgcggagc cacgtgggtg gtcctggggc actcagagag
      721 aaggcatgtc tttggggagt cagatgagct gattgggcag aaagtggccc atgctctggc
      781 agagggactc ggagtaatcg cctgcattgg ggagaagcta gatgaaaggg aagctggcat
      841 cactgagaag gttgttttcg agcagacaaa ggtcatcgca gataacgtga aggactggag
      901 caaggtcgtc ctggcctatg agcctgtgtg ggccgttggt actggcaaga ctgcaacacc
      961 ccaacaggcc caggaagtac acgagaagct ccgaggatgg ctgaagtcca acgtctctga
     1021 tgcggtggct cagagcaccc gtatcattta tggaggctct gtgactgggg caacctgcaa
     1081 ggagctggcc agccagcctg atgtggatgg cttccttgtg ggtggtgctt ccctcaagcc
     1141 cgaattcgtg gacatcatca atgccaaaca actcgagcac caccaccacc accactgaga
     1201 tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata
     1261 actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg
     1321 aactatatcc ggattggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt
     1381 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     1441 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     1501 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     1561 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     1621 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     1681 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     1741 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt
     1801 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     1861 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     1921 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     1981 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     2041 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     2101 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     2161 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     2221 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     2281 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     2341 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     2401 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     2461 gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     2521 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     2581 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     2641 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     2701 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     2761 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     2821 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     2881 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     2941 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     3001 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     3061 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     3121 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     3181 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     3241 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     3301 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     3361 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     3421 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     3481 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     3541 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     3601 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     3661 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     3721 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     3781 caccgcatat atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     3841 atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc
     3901 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac
     3961 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca
     4021 gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg cctgttcatc
     4081 cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc
     4141 catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct
     4201 gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga
     4261 tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg
     4321 gcgggaccag agaaaaatca
//
