LOCUS       sequence-97879-e        6316 bp    DNA     circular SYN 15-APR-2025
DEFINITION  to purify human ITPA from bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6316)
  AUTHORS   Porta J, Kolar C, Kozmin SG, Pavlov YI, Borgstahl GE
  TITLE     Structure of the orthorhombic form of human inosine triphosphate 
            pyrophosphatase.
  JOURNAL   Acta Crystallogr Sect F Struct Biol Cryst Commun. 2006 Nov 1;62(Pt 
            11):1076-81. Epub 2006 Oct 25.
  PUBMED    17077483
REFERENCE   2  (bases 1 to 6316)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Acta 
            Crystallogr Sect F Struct Biol Cryst Commun."; date: "2006-11-1"; 
            volume: "62(Pt 11)"; pages: "1076-81. Epub 2006 Oct 25"
COMMENT     Sequence Label: pET15b-ITPA
FEATURES             Location/Qualifiers
     source          1..6316
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..15,16..32,33..38)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      complement(213..260)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     256..274
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(942..959)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(969..986)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    1044..1068
                     /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(1068..1087)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1069..1087)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1171..1190)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1396..1473
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1474..2556
                     /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(1493..1512)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3868..4059
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4053..4075)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4161..4303
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4216..4235
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4318..4335)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4489..5077)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4569..4588)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5248..6039,6040..6108))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5871..5890
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6109..6213)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6281..6299
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ttctcatgtt tgacagctta tcatcgataa gctttaatgc ggtagtttat cacagttaaa
       61 ttgctaacgc agtcaggcac cgtgtatgaa atctaacaat gcgctcatcg tcatcctcgg
      121 caccgtcacc ctggatgctg taggcatagg cttggttatg ccggtactgc cgggcctctt
      181 gcgggatatc cggatatagt tcctcctttc agcaaaaaac ccctcaagac ccgtttagag
      241 gccccaaggg gttatgctag ttattgctca gcggtggcag cagccaactc agcttccttt
      301 cgggctttgt tagcagccgg atcctcgagc atatgctcct ccagctgcag aagtcaagct
      361 gccaaactgc caaagtactc ctgcagctcc agcagggccc ggaagcgatg ggagacagcg
      421 ttcttctccg ccttaggcat ctctgcgtac gtctgctcat atccatcagg ctgaaagcag
      481 gggtcccagc caaagtcctg gcagcctctg ggtgccacga tccggcccga ggtccggccc
      541 ctgaacaggc gcacgggctg gcttgggtcc ccggtgctga gtgcaaacgt gcagagcgca
      601 taggctgact tgtcctcgaa cccggccagg agctggtgga gaccttcagg ctttaacttc
      661 tccagaaacc actttatgta ggggccgggg agccctccaa gggcattgaa gcacagacaa
      721 gtgtcctcaa ccagcacggg cccctgtacc tggcgaactg cctcctgaca tttctgtatg
      781 gaaatctcat ccggctcccc ctggtactcc ggcaggtcaa ttttctgtgc caccaaagtg
      841 catggaaact tatctcctag aatctgaacg acctcctcca gcttcttggc gttccccgtt
      901 acaaacacga tcttcttccc caccaatgag gccgccatat ggctgccgcg cggcaccagg
      961 ccgctgctgt gatgatgatg atgatggctg ctgcccatgg tatatctcct tcttaaagtt
     1021 aaaccaaaat tattttctag aggggaattg ttatccgctc acaattcccc tatagtgagt
     1081 cgtattaatt tcgcgggatc gagatctcga tcctctacgc cggacgcatc gtggccggca
     1141 tcaccggcgc cacaggtgcg gttgctggcg cctatatcgc cgacatcacc gatggggaag
     1201 atcgggctcg ccacttcggg ctcatgagcg cttgtttcgg cgtgggtatg gtggcaggcc
     1261 ccgtggccgg gggactgttg ggcgccatct ccttgcatgc accattcctt gcggcggcgg
     1321 tgctcaacgg cctcaaccta ctactgggct gcttcctaat gcaggagtcg cataagggag
     1381 agcgtcgaga tcccggacac catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag
     1441 cgcccggaag agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc
     1501 gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac
     1561 gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc
     1621 aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc
     1681 agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa
     1741 ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg
     1801 gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat
     1861 gaccaggatg ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat
     1921 gtctctgacc agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg
     1981 ggcgtggagc atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta
     2041 agttctgtct cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa
     2101 attcagccga tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc
     2161 atgcaaatgc tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg
     2221 gcgctgggcg caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg
     2281 gtagtgggat acgacgatac cgaagacagc tcatgttata tcccgccgtt aaccaccatc
     2341 aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag
     2401 ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc
     2461 ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     2521 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta
     2581 gctcactcat taggcaccgg gatctcgacc gatgcccttg agagccttca acccagtcag
     2641 ctccttccgg tgggcgcggg gcatgactat cgtcgccgca cttatgactg tcttctttat
     2701 catgcaactc gtaggacagg tgccggcagc gctctgggtc attttcggcg aggaccgctt
     2761 tcgctggagc gcgacgatga tcggcctgtc gcttgcggta ttcggaatct tgcacgccct
     2821 cgctcaagcc ttcgtcactg gtcccgccac caaacgtttc ggcgagaagc aggccattat
     2881 cgccggcatg gcggccgacg cgctgggcta cgtcttgctg gcgttcgcga cgcgaggctg
     2941 gatggccttc cccattatga ttcttctcgc ttccggcggc atcgggatgc ccgcgttgca
     3001 ggccatgctg tccaggcagg tagatgacga ccatcaggga cagcttcaag gatcgctcgc
     3061 ggctcttacc agcctaactt cgatcactgg accgctgatc gtcacggcga tttatgccgc
     3121 ctcggcgagc acatggaacg ggttggcatg gattgtaggc gccgccctat accttgtctg
     3181 cctccccgcg ttgcgtcgcg gtgcatggag ccgggccacc tcgacctgaa tggaagccgg
     3241 cggcacctcg ctaacggatt caccactcca agaattggag ccaatcaatt cttgcggaga
     3301 actgtgaatg cgcaaaccaa cccttggcag aacatatcca tcgcgtccgc catctccagc
     3361 agccgcacgc ggcgcatctc gggcagcgtt gggtcctggc cacgggtgcg catgatcgtg
     3421 ctcctgtcgt tgaggacccg gctaggctgg cggggttgcc ttactggtta gcagaatgaa
     3481 tcaccgatac gcgagcgaac gtgaagcgac tgctgctgca aaacgtctgc gacctgagca
     3541 acaacatgaa tggtcttcgg tttccgtgtt tcgtaaagtc tggaaacgcg gaagtcagcg
     3601 ccctgcacca ttatgttccg gatctgcatc gcaggatgct gctggctacc ctgtggaaca
     3661 cctacatctg tattaacgaa gcgctggcat tgaccctgag tgatttttct ctggtcccgc
     3721 cgcatccata ccgccagttg tttaccctca caacgttcca gtaaccgggc atgttcatca
     3781 tcagtaaccc gtatcgtgag catcctctct cgtttcatcg gtatcattac ccccatgaac
     3841 agaaatcccc cttacacgga ggcatcagtg accaaacagg aaaaaaccgc ccttaacatg
     3901 gcccgcttta tcagaagcca gacattaacg cttctggaga aactcaacga gctggacgcg
     3961 gatgaacagg cagacatctg tgaatcgctt cacgaccacg ctgatgagct ttaccgcagc
     4021 tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     4081 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     4141 ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat
     4201 actggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat atatgcggtg
     4261 tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt ccgcttcctc
     4321 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     4381 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     4441 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     4501 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     4561 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     4621 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     4681 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     4741 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     4801 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     4861 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     4921 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     4981 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     5041 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     5101 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     5161 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     5221 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     5281 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     5341 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     5401 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     5461 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     5521 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc
     5581 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     5641 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     5701 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     5761 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     5821 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaacacggg ataataccgc
     5881 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     5941 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     6001 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     6061 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     6121 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     6181 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     6241 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     6301 ctttcgtctt caagaa
//
