LOCUS       sequence-190308-        6261 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human PPAT-EGFP fusion in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6261)
  AUTHORS   An S, Kumar R, Sheets ED, Benkovic SJ
  TITLE     Reversible compartmentalization of de novo purine biosynthetic 
            complexes in living cells.
  JOURNAL   Science. 2008 Apr 4;320(5872):103-6. doi: 10.1126/science.1152241.
  PUBMED    18388293
REFERENCE   2  (bases 1 to 6261)
  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: "10.1126/science"; 
            journalName: "Science"; date: "2008-04-4- 4"; volume: "320"; issue: 
            "5872"; pages: "103-6"
COMMENT     Sequence Label: pPPAT-EGFP
FEATURES             Location/Qualifiers
     source          1..6261
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2201..2210
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2207..2209,2210..2212,2213..2926)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLK
                     ERVTAAGITLGMDELYK"
     primer_bind     complement(2252..2273)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2513..2532)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    3049..3170
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3086..3105)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3140..3159
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3177..3632)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3314..3335)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3526..3545
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3659..3763
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3765..4122
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3765..3956
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3788..3808)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3973..4108
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4157..4951
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4157..4951
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4211..4230)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4821..4840
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5139..5158)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5183..5230
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5559..6147
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6048..6067
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagcc atatggagct ggaggagttg gggatccgag aggaatgtgg
      661 cgtgttcggg tgcatcgcct caggagagtg gcccacgcag ctggatgtac cgcatgtgat
      721 cactctggga ctcgtggggc tgcagcaccg gggtcaggag agtgctggta ttgtgactag
      781 tgatgggagt tccgtgccaa cattcaaatc acacaaggga atgggtcttg taaatcacgt
      841 ctttactgaa gacaatttga aaaaattata tgtttcaaat cttggaattg gacacaccag
      901 gtatgccacc acaggaaaat gtgaactaga aaattgtcag cccttcgttg ttgaaacact
      961 tcatgggaag atagctgtgg cacataatgg cgaattggta aatgctgctc gattaaggaa
     1021 aaagcttctg cgtcatggta ttggtctgtc tacaagttct gatagtgaaa tgattaccca
     1081 gttactggcg tatacccctc ctcaggaaca agatgacacc ccagactggg tagccaggat
     1141 taaaaacttg atgaaggaag cacccacagc atactccctg cttataatgc acagagatgt
     1201 tatttatgca gtacgagatc cttatggaaa tcgtccctta tgcattggtc gtcttattcc
     1261 agtgtctgat ataaatgaca aagagaaaaa aacatcagaa acagaaggat gggtggtgtc
     1321 ttcagaatct tgtagcttct tatctattgg tgcaagatat taccgtgaag tcttgcctgg
     1381 agaaattgtg gaaatatcca gacacaatgt ccaaactctt gatattatat caaggtctga
     1441 aggaaaccca gtggcttttt gtatctttga atatgtttat tttgcaagac cagacagtat
     1501 gttcgaagac caaatggttt atacagtaag ataccgttgt ggccagcagc tagcgattga
     1561 agcacctgtg gatgcagatt tggttagcac tgttccagaa tctgctacgc ctgctgctct
     1621 tgcttacgca ggaaagtgtg gacttccata tgtggaggtg ctgtgtaaaa accggtatgt
     1681 agggagaacc ttcattcagc caaacatgag gttaagacaa cttggtgttg caaaaaaatt
     1741 tggagtattg tcagacaact ttaaaggcaa aagaattgtt cttgtagatg attcaattgt
     1801 cagaggcaat accatctcac ctataataaa actgctcaaa gaatctggtg caaaagaggt
     1861 acacattcga gtagcttcac caccaattaa atatccatgc ttcatgggaa taaacattcc
     1921 tacaaaagaa gagctcattg ccaataaacc agaatttgat caccttgcag aatatctagg
     1981 agcaaacagt gttgtgtatc tgtcagtaga aggactggtt tcatctgtac aagaagggat
     2041 aaagtttaaa aaacagaaag agaaaaagca cgatattatg atccaagaaa atggaaatgg
     2101 tctggaatgt tttgaaaaga gtggtcattg tacagcttgt ctcactggaa aatatcctgt
     2161 agaattagaa tgggtaccgc gggcccggga tccaccggtc gccaccatgg tgagcaaggg
     2221 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     2281 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     2341 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
     2401 gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt
     2461 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     2521 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     2581 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     2641 ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa
     2701 cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca
     2761 gaacaccccc atcggcgacg gtcccgtgct gctgcccgac aaccactacc tgagcaccca
     2821 gtccaagctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctga aggagcgcgt
     2881 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtaaagcg gccgcgactc
     2941 tagatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     3001 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     3061 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     3121 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta aggcgtaaat
     3181 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     3241 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     3301 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     3361 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     3421 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     3481 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     3541 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     3601 cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg
     3661 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     3721 ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac
     3781 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3841 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3901 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3961 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     4021 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     4081 aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg
     4141 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     4201 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     4261 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     4321 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     4381 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     4441 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     4501 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     4561 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     4621 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     4681 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     4741 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     4801 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     4861 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     4921 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     4981 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     5041 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     5101 atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata
     5161 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg
     5221 tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg
     5281 agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt
     5341 tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca
     5401 ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     5461 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     5521 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     5581 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     5641 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     5701 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     5761 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     5821 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     5881 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     5941 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     6001 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     6061 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     6121 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     6181 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     6241 aaccgtatta ccgccatgca t
//
