LOCUS       sequence-330489-        6250 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human PPAT-mCherry fusion in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6250)
  AUTHORS   Pedley AM, Benkovic SJ
  TITLE     Detecting Purinosome Metabolon Formation with Fluorescence 
            Microscopy.
  JOURNAL   Methods Mol Biol. 2018;1764:279-289. doi: 
            10.1007/978-1-4939-7759-8_17.
  PUBMED    29605921
REFERENCE   2  (bases 1 to 6250)
  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: "Methods Mol
            Biol."; date: "2018"; volume: "1764:279-289. doi"; pages: " 
            10.1007/978-1-4939-7759-8_17"
COMMENT     Sequence Label: pPPAT-mCherry
FEATURES             Location/Qualifiers
     source          1..6250
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2141..2150
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2147..2857
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2292..2310)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2560..2579
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2978..3099
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3015..3034)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3069..3088
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3106..3561)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3243..3264)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3455..3474
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3588..3692
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3694..4051
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3694..3885
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3717..3737)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3902..4037
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4086..4880
                     /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             4086..4880
                     /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(4140..4159)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4750..4769
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5068..5087)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5112..5159
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5488..6076
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5977..5996
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggactcag atctcgagcc atatggagct ggaggagttg gggatccgag aggaatgtgg
      601 cgtgttcggg tgcatcgcct caggagagtg gcccacgcag ctggatgtac cgcatgtgat
      661 cactctggga ctcgtggggc tgcagcaccg gggtcaggag agtgctggta ttgtgactag
      721 tgatgggagt tcggtgccaa cattcaaatc acacaaggga atgggtcttg taaatcacgt
      781 ctttactgaa gacaatttga aaaaattata tgtttcaaat cttggaattg gacacaccag
      841 gtatgccacc acaggaaaat gtgaactaga aaattgtcag cccttcgttg ttgaaacact
      901 tcatgggaag atagctgtgg cacataatgg cgaattggta aatgctgctc gattaaggaa
      961 aaagcttctg cgtcatggta ttggtctgtc tacaagttct gatagtgaaa tgattaccca
     1021 gttactggcg tatacccctc ctcaggaaca agatgacacc ccagactggg tagccaggat
     1081 taaaaacttg atgaaggaag cacccacagc atactccctg cttataatgc acagagatgt
     1141 tatttatgca gtacgagatc cttatggaaa tcgtccctta tgcattggtc gtcttattcc
     1201 agtgtctgat ataaatgaca aagagaaaaa aacatcagaa acagaaggat gggtggtgtc
     1261 ttcagaatct tgtagcttct tatctattgg tgcaagatat taccgtgaag tcttgcctgg
     1321 agaaattgtg gaaatatcca gacacaatgt ccaaactctt gatattatat caaggtctga
     1381 aggaaaccca gtggcttttt gtatctttga atatgtttat tttgcaagac cagacagtat
     1441 gttcgaagac caaatggttt atacagtaag ataccgttgt ggccagcagc tagcgattga
     1501 agcacctgtg gatgcagatt tggttagcac tgttccagaa tctgctacgc ctgctgctct
     1561 tgcttacgca ggaaagtgtg gacttccata tgtggaggtg ctgtgtaaaa accggtatgt
     1621 agggagaacc ttcattcagc caaacatgag gttaagacaa cttggtgttg caaaaaaatt
     1681 tggagtattg tcagacaact ttaaaggcaa aagaattgtt cttgtagatg attcaattgt
     1741 cagaggcaat accatctcac ctataataaa actgctcaaa gaatctggtg caaaagaggt
     1801 acacattcga gtagcttcac caccaattaa atatccatgc ttcatgggaa taaacattcc
     1861 tacaaaagaa gagctcattg ccaataaacc agaatttgat caccttgcag aatatctagg
     1921 agcaaacagt gttgtgtatc tgtcagtaga aggactggtt tcatctgtac aagaagggat
     1981 aaagtttaaa aaacagaaag agaaaaagca cgatattatg atccaagaaa atggaaatgg
     2041 tctggaatgt tttgaaaaga gtggtcattg tacagcttgt ctcactggaa aatatcctgt
     2101 agaattagaa tgggtaccgc gggcccggga tccaccggtc gccaccatgg tgagcaaggg
     2161 cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg
     2221 ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg
     2281 cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat
     2341 cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc
     2401 cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga
     2461 ggacggcggc gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta
     2521 caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac
     2581 catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga
     2641 gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac
     2701 ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga
     2761 catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg
     2821 ccactccacc ggcggcatgg acgagctgta caagtagcgg ccgcgactct agatcataat
     2881 cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac acctccccct
     2941 gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg cagcttataa
     3001 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     3061 ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa ggcgtaaatt gtaagcgtta
     3121 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     3181 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     3241 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
     3301 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg
     3361 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
     3421 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
     3481 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
     3541 atgcgccgct acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     3601 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     3661 aaatgcttca ataatattga aaaaggaaga gtcctgaggc ggaaagaacc agctgtggaa
     3721 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     3781 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     3841 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     3901 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     3961 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     4021 aggctttttt ggaggcctag gcttttgcaa agatcgatca agagacagga tgaggatcgt
     4081 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     4141 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     4201 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     4261 aactgcaaga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     4321 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     4381 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     4441 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     4501 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     4561 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgagcatgc
     4621 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     4681 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     4741 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     4801 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     4861 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     4921 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     4981 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     5041 cttcgcccac cctaggggga ggctaactga aacacggaag gagacaatac cggaaggaac
     5101 ccgcgctatg acggcaataa aaagacagaa taaaacgcac ggtgttgggt cgtttgttca
     5161 taaacgcggg gttcggtccc agggctggca ctctgtcgat accccaccga gaccccattg
     5221 gggccaatac gcccgcgttt cttccttttc cccaccccac cccccaagtt cgggtgaagg
     5281 cccagggctc gcagccaacg tcggggcggc aggccctgcc atagcctcag gttactcata
     5341 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     5401 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     5461 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     5521 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     5581 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     5641 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     5701 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     5761 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     5821 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     5881 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     5941 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     6001 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     6061 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     6121 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     6181 cgccatgcat tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     6241 tggagttccg
//
