LOCUS       sequence-190228-        6242 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  hQPRTase harbouring K139A mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6242)
  AUTHORS   Liu H, Woznica K, Catton G, Crawford A, Botting N, Naismith JH
  TITLE     Structural and kinetic characterization of quinolinate 
            phosphoribosyltransferase (hQPRTase) from homo sapiens.
  JOURNAL   J Mol Biol. 2007 Oct 26;373(3):755-63. Epub 2007 Aug 24.
  PUBMED    17868694
REFERENCE   2  (bases 1 to 6242)
  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: "J Mol 
            Biol."; date: "2007-10-26"; volume: "373(3)"; pages: "755-63. Epub 
            2007 Aug 24"
COMMENT     Sequence Label: pEHIShqprtK139A
FEATURES             Location/Qualifiers
     source          1..6242
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1100..1120)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(1142..1159)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1179..1201
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1216..1240
                     /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)."
     primer_bind     complement(1240..1259)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1241..1259)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1347..1366)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1572..1649
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1650..2732
                     /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     complement(1669..1688)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3541..3732
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3726..3748)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3834..3973
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3889..3908
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3988..4005)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4159..4747)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4239..4258)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4858..4877)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4869..5684
                     /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             4869..5684
                     /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     complement(4942..4961)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5776..6231)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5913..5934)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6125..6144
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tcctagtgga ttttgggcac tggagccacc tctttggcaa
      241 acagcttgag ggagaaatca agggctgggg ccgcctgggt cagcatcccc atggagatga
      301 cgtctatgtg cggcccgcag aactggggga ggttgtccag ggtgatgccc ccactggctt
      361 ccacagccac actcgggaac tgggccttca gcacggtggc cgtggggtgc agctcctctg
      421 gcttgaagtt gtccagcagg acaaggtcgg caccagcctc agctgcctgc acggcctcct
      481 gcaggctgct gcattccact tccaccttca gagcgaagtc agccgcctgt ctggccgccc
      541 gcaccgcctt ctccacgcca ccggcggcca ccacatggtt atccttcacc atcaccagcc
      601 ctcccaggtc gtagcggtgc gaggcggccc cgcccaccag gagcccatac ttctccacca
      661 gccggaagcc tggcgtggtc gccctcgtgc ctgccacgtg cccagtccag ccggcccccc
      721 tggcggcctc cactgcagcg gcggcagcac tggcaatgcc actgcagcgg gccagcgtgt
      781 tgagggccac ccgttccccc agcagcaggc agtgggcagg gccccggacc tcggccactc
      841 tggccaccgg caccagcttc gatccctcgg ggaggaacca ggagacttgg cagttgagtt
      901 gggtaaatat ggcatcgaag aaaggctgcc ctgccagtac cccaggggat ttggcccaca
      961 gcgccgcctg cgaggggcct gccccgctga ccaaggctgc gtagttgagc cctgggcagt
     1021 cctctcggag ccagctgtcc accagggctg ccagggtgac gggcggcagc agcagcgcca
     1081 ggccttcagc gttcatggcg ccctgaaaat acaggttttc ggtcgttggg atatcgtaat
     1141 cgtgatggtg atggtgatgg tagtacgaca tatgtatatc tccttcttaa agttaaacaa
     1201 aattatttct agaggggaat tgttatccgc tcacaattcc cctatagtga gtcgtattaa
     1261 tttcgcggga tcgagatcga tctcgatcct ctacgccgga cgcatcgtgg ccggcatcac
     1321 cggcgccaca ggtgcggttg ctggcgccta tatcgccgac atcaccgatg gggaagatcg
     1381 ggctcgccac ttcgggctca tgagcgcttg tttcggcgtg ggtatggtgg caggccccgt
     1441 ggccggggga ctgttgggcg ccatctcctt gcatgcacca ttccttgcgg cggcggtgct
     1501 caacggcctc aacctactac tgggctgctt cctaatgcag gagtcgcata agggagagcg
     1561 tcgagatccc ggacaccatc gaatggcgca aaacctttcg cggtatggca tgatagcgcc
     1621 cggaagagag tcaattcagg gtggtgaatg tgaaaccagt aacgttatac gatgtcgcag
     1681 agtatgccgg tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc agccacgttt
     1741 ctgcgaaaac gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac attcccaacc
     1801 gcgtggcaca acaactggcg ggcaaacagt cgttgctgat tggcgttgcc acctccagtc
     1861 tggccctgca cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc gatcaactgg
     1921 gtgccagcgt ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt aaagcggcgg
     1981 tgcacaatct tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg ctggatgacc
     2041 aggatgccat tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt cttgatgtct
     2101 ctgaccagac acccatcaac agtattattt tctcccatga agacggtacg cgactgggcg
     2161 tggagcatct ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt
     2221 ctgtctcggc gcgtctgcgt ctggctggct ggcataaata tctcactcgc aatcaaattc
     2281 agccgatagc ggaacgggaa ggcgactgga gtgccatgtc cggttttcaa caaaccatgc
     2341 aaatgctgaa tgagggcatc gttcccactg cgatgctggt tgccaacgat cagatggcgc
     2401 tgggcgcaat gcgcgccatt accgagtccg ggctgcgcgt tggtgcggac atctcggtag
     2461 tgggatacga cgataccgaa gacagctcat gttatatccc gccgttaacc accatcaaac
     2521 aggattttcg cctgctgggg caaaccagcg tggaccgctt gctgcaactc tctcagggcc
     2581 aggcggtgaa gggcaatcag ctgttgcccg tctcactggt gaaaagaaaa accaccctgg
     2641 cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac
     2701 gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt aagttagctc
     2761 actcattagg caccgggatc tcgaccgatg cccttgagag ccttcaaccc agtcagctcc
     2821 ttccggtggg cgcggggcat gactatcgtc gccgcactta tgactgtctt ctttatcatg
     2881 caactcgtag gacaggtgcc ggcagcgctc tgggtcattt tcggcgagga ccgctttcgc
     2941 tggagcgcga cgatgatcgg cctgtcgctt gcggtattcg gaatcttgca cgccctcgct
     3001 caagccttcg tcactggtcc cgccaccaaa cgtttcggcg agaagcaggc cattatcgcc
     3061 ggcatggcgg ccccacgggt gcgcatgatc gtgctcctgt cgttgaggac ccggctaggc
     3121 tggcggggtt gccttactgg ttagcagaat gaatcaccga tacgcgagcg aacgtgaagc
     3181 gactgctgct gcaaaacgtc tgcgacctga gcaacaacat gaatggtctt cggtttccgt
     3241 gtttcgtaaa gtctggaaac gcggaagtca gcgccctgca ccattatgtt ccggatctgc
     3301 atcgcaggat gctgctggct accctgtgga acacctacat ctgtattaac gaagcgctgg
     3361 cattgaccct gagtgatttt tctctggtcc cgccgcatcc ataccgccag ttgtttaccc
     3421 tcacaacgtt ccagtaaccg ggcatgttca tcatcagtaa cccgtatcgt gagcatcctc
     3481 tctcgtttca tcggtatcat tacccccatg aacagaaatc ccccttacac ggaggcatca
     3541 gtgaccaaac aggaaaaaac cgcccttaac atggcccgct ttatcagaag ccagacatta
     3601 acgcttctgg agaaactcaa cgagctggac gcggatgaac aggcagacat ctgtgaatcg
     3661 cttcacgacc acgctgatga gctttaccgc agctgcctcg cgcgtttcgg tgatgacggt
     3721 gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc
     3781 gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc
     3841 atgacccagt cacgtagcga tagcggagtg tatactggct taactatgcg gcatcagagc
     3901 agattgtact gagagtgcac cattgcggtg tgaaataccg cacagatgcg taaggagaaa
     3961 ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     4021 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     4081 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     4141 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     4201 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     4261 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     4321 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     4381 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     4441 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     4501 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     4561 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     4621 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     4681 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     4741 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     4801 acgttaaggg attttggtca tgaacaataa aactgtctgc ttacataaac agtaatacaa
     4861 ggggtgttat gagccatatt caacgggaaa cgtcttgctc taggccgcga ttaaattcca
     4921 acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg caatcaggtg
     4981 cgacaatcta tcgattgtat gggaagcccg atgcgccaga gttgtttctg aaacatggca
     5041 aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg ctgacggaat
     5101 ttatgcctct tccgaccatc aagcatttta tccgtactcc tgatgatgca tggttactca
     5161 ccactgcgat ccccgggaaa acagcattcc aggtattaga agaatatcct gattcaggtg
     5221 aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta
     5281 attgtccttt taacagcgat cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata
     5341 acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct gttgaacaag
     5401 tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc actcatggtg
     5461 atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt attgatgttg
     5521 gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac tgcctcggtg
     5581 agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat aatcctgata
     5641 tgaataaatt gcagtttcat ttgatgctcg atgagttttt ctaagaatta attcatgagc
     5701 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     5761 cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     5821 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     5881 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     5941 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     6001 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     6061 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     6121 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     6181 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc
     6241 ca
//
