LOCUS       sequence-245695-        7065 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses human PPAT protein, with first 11 residues removed, with a
            C-terminal 2XStrep affinity tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7065)
  AUTHORS   Liu C, Knudsen GM, Pedley AM, He J, Johnson JL, Yaron TM, Cantley 
            LC, Benkovic SJ
  TITLE     Mapping Post-Translational Modifications of de Novo Purine 
            Biosynthetic Enzymes: Implications for Pathway Regulation.
  JOURNAL   J Proteome Res. 2019 May 3;18(5):2078-2087. doi: 
            10.1021/acs.jproteome.8b00969. Epub 2019 Apr 18.
  PUBMED    30964683
REFERENCE   2  (bases 1 to 7065)
  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"; doi: 
            "10.1021/acs.jproteome.8b00969"; journalName: "J Proteome Res"; 
            date: "2019-05-3- 3"; volume: "18"; issue: "5"; pages: "2078-2087"
COMMENT     Sequence Label: pPPAT-11AA-2XStrep
FEATURES             Location/Qualifiers
     source          1..7065
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..729
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     680..700
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     726..750
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          890..1022
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1005..1024
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1067..1085
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2627..2632,2633..2656,2657..2692,2693..2716)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="SAWSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     polyA_signal    2769..2890
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2780..2799)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2834..2853
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      3076..3531
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3163..3182)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3373..3394
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3648..4005
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3648..3839
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3671..3691)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3856..3991
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4056..4850
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4110..4129)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4720..4739
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4914..4962
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(5003..5022)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5122..5144
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5182..5200)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5268..5372
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5373..5441,5442..6233)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5591..5610)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6404..6992
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6893..6912
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactg
      661 cgatcgcccg ccccgttgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
      721 agcagagctc gtttagtgaa ccgtcagatc actagaagct ttattgcggt agtttatcac
      781 agttaaattg ctaacgcagt cagtgcttct gacacaacag tctcgaactt aagctgcagt
      841 gactctctta aggtagcctt gcagaagttg gtcgtgaggc actgggcagg taagtatcaa
      901 ggttacaaga caggtttaag gagaccaata gaaactgggc ttgtcgagac agagaagact
      961 cttgcgtttc tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac
     1021 aggtgtccac tcccagttca attacagctc ttaaggctag agtacttaat acgactcact
     1081 ataggctagc ctcgagatgt gtggcgtgtt cgggtgcatc gcctcaggag agtggcccac
     1141 gcagctggat gtaccgcatg tgatcactct gggactcgtg gggctgcagc accggggtca
     1201 ggagagtgct ggtattgtga ctagtgatgg gagttccgtg ccaacattca aatcacacaa
     1261 gggaatgggt cttgtaaatc acgtctttac tgaagacaat ttgaaaaaat tatatgtttc
     1321 aaatcttgga attggacaca ccaggtatgc caccacagga aaatgtgaac tagaaaattg
     1381 tcagcccttc gttgttgaaa cacttcatgg gaagatagct gtggcacata atggcgaatt
     1441 ggtaaatgct gctcgattaa ggaaaaagct tctgcgtcat ggtattggtc tgtctacaag
     1501 ttctgatagt gaaatgatta cccagttact ggcgtatacc cctcctcagg aacaagatga
     1561 caccccagac tgggtagcca ggattaaaaa cttgatgaag gaagcaccca cagcatactc
     1621 cctgcttata atgcacagag atgttattta tgcagtacga gatccttatg gaaatcgtcc
     1681 cttatgcatt ggtcgtctta ttccagtgtc tgatataaat gacaaagaga aaaaaacatc
     1741 agaaacagaa ggatgggtgg tgtcttcaga atcttgtagc ttcttatcta ttggtgcaag
     1801 atattaccgt gaagtcttgc ctggagaaat tgtggaaata tccagacaca atgtccaaac
     1861 tcttgatatt atatcaaggt ctgaaggaaa cccagtggct ttttgtatct ttgaatatgt
     1921 ttattttgca agaccagaca gtatgttcga agaccaaatg gtttatacag taagataccg
     1981 ttgtggccag cagctagcga ttgaagcacc tgtggatgca gatttggtta gcactgttcc
     2041 agaatctgct acgcctgctg ctcttgctta cgcaggaaag tgtggacttc catatgtgga
     2101 ggtgctgtgt aaaaaccggt atgtagggag aaccttcatt cagccaaaca tgaggttaag
     2161 acaacttggt gttgcaaaaa aatttggagt attgtcagac aactttaaag gcaaaagaat
     2221 tgttcttgta gatgattcaa ttgtcagagg caataccatc tcacctataa taaaactgct
     2281 caaagaatct ggtgcaaaag aggtacacat tcgagtagct tcaccaccaa ttaaatatcc
     2341 atgcttcatg ggaataaaca ttcctacaaa agaagagctc attgccaata aaccagaatt
     2401 tgatcacctt gcagaatatc taggagcaaa cagtgttgtg tatctgtcag tagaaggact
     2461 ggtttcatct gtacaagaag ggataaagtt taaaaaacag aaagagaaaa agcacgatat
     2521 tatgatccaa gaaaatggaa atggtctgga atgttttgaa aagagtggtc attgtacagc
     2581 ttgtctcact ggaaaatatc ctgtagaatt agaatggata cccgggagcg cttggagcca
     2641 cccgcagttc gaaaaaggtg gaggttctgg cggtggatcg ggaggttcag cgtggagcca
     2701 cccgcagttc gagaaataaa gcgcggccgc ttccctttag tgagggttaa tgcttcgagc
     2761 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     2821 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     2881 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggagatgtg
     2941 ggaggttttt taaagcaagt aaaacctcta caaatgtggt aaaatccgat aaggatcgat
     3001 ccgggctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     3061 ctgaatggcg aatggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     3121 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     3181 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     3241 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     3301 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     3361 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     3421 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     3481 tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt tcctgatgcg
     3541 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgcggat ctgcgcagca
     3601 ccatggcctg aaataacctc tgaaagagga acttggttag gtaccttctg aggcggaaag
     3661 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     3721 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     3781 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     3841 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     3901 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     3961 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct tgattcttct
     4021 gacacaacag tctcgaactt aaggctagag ccaccatgat tgaacaagat ggattgcacg
     4081 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     4141 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     4201 tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt
     4261 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     4321 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     4381 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     4441 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     4501 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     4561 aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg
     4621 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     4681 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     4741 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     4801 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     4861 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgatggc cgcaataaaa
     4921 tatctttatt ttcattacat ctgtgtgttg gttttttgtg tgaatcgata gcgataagga
     4981 tccgcgtatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     5041 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     5101 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     5161 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     5221 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     5281 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     5341 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     5401 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     5461 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     5521 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     5581 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     5641 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     5701 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     5761 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     5821 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     5881 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     5941 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     6001 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     6061 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     6121 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     6181 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     6241 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     6301 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     6361 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     6421 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     6481 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     6541 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     6601 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     6661 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     6721 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     6781 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     6841 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     6901 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     6961 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     7021 tcctggcctt ttgctggcct tttgctcaca tggctcgaca gatct
//
