LOCUS       sequence-236915-        8479 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Produces equimolar expression of yeast codon-optimized mTq2 and 
            mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8479)
  AUTHORS   Botman D, de Groot DH, Schmidt P, Goedhart J, Teusink B
  TITLE     In vivo characterisation of fluorescent proteins in budding yeast.
  JOURNAL   Sci Rep. 2019 Feb 19;9(1):2234. doi: 10.1038/s41598-019-38913-z.
  PUBMED    30783202
REFERENCE   2  (bases 1 to 8479)
  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: "Sci Rep."; 
            date: "2019-02-19"; volume: "9(1):2234. doi"; pages: " 
            10.1038/s41598-019-38913-z"
COMMENT     Sequence Label: pDRF1-GW ymTurquoise2-T2A-mCherry
FEATURES             Location/Qualifiers
     source          1..8479
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    800..824
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1574..1627
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     regulatory      1643..1652
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1649..2359
                     /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(1794..1812)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2062..2081
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(2369..2393)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2455..2474)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2456..2474)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      2622..2809
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2677..2701)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(2821..2838)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2821..2837)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2830..2852)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3273..4619
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     CDS             complement(4730..5533)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(5534..5749)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(5774..5792)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5860..5964
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5965..6033,6034..6825)
                     /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(6183..6202)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6996..7584
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7485..7504
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7738..7755
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7872..7893
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7908..7913,7914..7931,7932..7938)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7946..7962
                     /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     7951..7973
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7970..7986
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7970..7986
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     rep_origin      complement(8016..8471)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8153..8174)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8365..8384
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 caagcttcct gaaacggaga aacataaaca ggcattgctg ggatcaccca tacatcactc
       61 tgttttgcct gaccttttcc ggtaatttga aaacaaaccc ggtctcgaag cggagatccg
      121 gcgataatta ccgcagaaat aaacccatac acgagacgta gaaccagccg cacatggccg
      181 gagaaactcc tgcgagaatt tcgtaaactc gcgcgcattg catctgtatt tcctaatgcg
      241 gcacttccag gcctcgatcg agaccgttta tccattgctt ttttgttgtc tttttccctc
      301 gttcacagaa agtctgaaga agctatagta gaactatgag ctttttttgt ttctgttttc
      361 cttttttttt tttttacctc tgtggaaatt gttactctca cactctttag ttcgtttgtt
      421 tgttttgttt attccaatta tgaccggtga cgaaacgtgg tcgatggtgg gtaccgctta
      481 tgctcccctc cattagtttc gattatataa aaaggccaaa tattgtatta ttttcaaatg
      541 tcctatcatt atcgtctaac atctaatttc tcttaaattt tttctctttc tttcctataa
      601 caccaatagt gaaaatcttt ttttcttcta tatctacaaa aacttttttt ttctatcaac
      661 ctcgttgata aattttttct ttaacaatcg ttaataatta attaattgga aaataaccat
      721 tttttctctc ttttatacac acattcaaaa gaaagaaaaa aaatataccc cagcctcgac
      781 tagtgaattc gcggccatca caagtttgta caaaaaagca ggctgctagc actagtaagc
      841 ttttaattaa atggttagta aaggtgaaga attgttcact ggtgtagtgc caattttagt
      901 agaattagat ggtgacgtta acggtcataa gttctctgta agcggtgaag gtgaaggtga
      961 cgcaacatac ggtaaattaa ctttgaaatt catttgtact acaggtaaat taccagttcc
     1021 atggccaact ttggttacta ctttgtcttg gggtgttcaa tgtttcgcta gatacccaga
     1081 tcatatgaag caacatgatt tctttaagtc tgctatgcca gaaggttacg ttcaagaaag
     1141 aactatcttc ttcaaggatg atggtaacta taagactaga gctgaagtta agttcgaagg
     1201 agatacttta gttaatagaa ttgaattgaa gggtattgat tttaaggaag atggtaatat
     1261 tttgggtcat aagttggaat ataattactt ctctgataac gtttacatta ctgctgataa
     1321 gcaaaagaac ggtattaagg cgaacttcaa gattagacat aacattgaag atggtggtgt
     1381 tcaattagct gatcattacc aacaaaacac tccaattggt gacggtccag ttttgttgcc
     1441 agataaccat tacttgtcta ctcaatctaa gttgtctaag gatccaaacg aaaagagaga
     1501 tcatatggtt ttgttggaat tcgttactgc tgctgggatc actttaggta tggatgaatt
     1561 gtataaatcc ggagagggca gaggaagtct tctaacatgc ggtgacgtgg aggagaatcc
     1621 cggccctggg gatccaccgg tcgccaccat ggtgagcaag ggcgaggagg ataacatggc
     1681 catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga acggccacga
     1741 gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct
     1801 gaaggtgacc aagggtggcc ccctgccctt cgcctgggac atcctgtccc ctcagttcat
     1861 gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact tgaagctgtc
     1921 cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac
     1981 cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg
     2041 caccaacttc ccctccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc
     2101 ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc agaggctgaa
     2161 gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg ccaagaagcc
     2221 cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct cccacaacga
     2281 ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca ccggcggcat
     2341 ggacgagctg tacaagtaaa gcggccgcac ccagctttct tgtacaaagt ggtgtagtta
     2401 attcatgatg gccgctgcag gtcgacctcg agggggggcc cggtacccaa ttcgccctat
     2461 agtgagtcgt attacgcgcg gatccagctt tggacttctt cgccagaggt ttggtcaagt
     2521 ctccaatcaa ggttgtcggc ttgtctacct tgccagaaat ttacgaaaag atggaaaagg
     2581 gtcaaatcgt tggtagatac gttgttgaca cttctaaata agcgaatttc ttatgattta
     2641 tgatttttat tattaaataa gttataaaaa aaataagtgt atacaaattt taaagtgact
     2701 cttaggtttt aaaacgaaaa ttcttattct tgagtaactc tttcctgtag gtcaggttgc
     2761 tttctcaggt atagcatgag gtcgctctta ttgaccacac ctctaccggc atgccaattc
     2821 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
     2881 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
     2941 cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt attttctcct
     3001 tacgcatctg tgcggtattt cacaccgcat aatcggatcg tacttgttac ccatcattga
     3061 attttgaaca tccgaacctg ggagttttcc ctgaaacaga tagtatattt gaacctgtat
     3121 aataatatat agtctagcgc tttacggaag acaatgtatg tatttcggtt cctggagaaa
     3181 ctattgcatc tattgcatag gtaatcttgc acgtcgcatc cccggttcat tttctgcgtt
     3241 tccatcttgc acttcaatag catatctttg ttaacgaagc atctgtgctt cattttgtag
     3301 aacaaaaatg caacgcgaga gcgctaattt ttcaaacaaa gaatctgagc tgcattttta
     3361 cagaacagaa atgcaacgcg aaagcgctat tttaccaacg aagaatctgt gcttcatttt
     3421 tgtaaaacaa aaatgcaacg cgagagcgct aatttttcaa acaaagaatc tgagctgcat
     3481 ttttacagaa cagaaatgca acgcgagagc gctattttac caacaaagaa tctatacttc
     3541 ttttttgttc tacaaaaatg catcccgaga gcgctatttt tctaacaaag catcttagat
     3601 tacttttttt ctcctttgtg cgctctataa tgcagtctct tgataacttt ttgcactgta
     3661 ggtccgttaa ggttagaaga aggctacttt ggtgtctatt ttctcttcca taaaaaaagc
     3721 ctgactccac ttcccgcgtt tactgattac tagcgaagct gcgggtgcat tttttcaaga
     3781 taaaggcatc cccgattata ttctataccg atgtggattg cgcatacttt gtgaacagaa
     3841 agtgatagcg ttgatgattc ttcattggtc agaaaattat gaacggtttc ttctattttg
     3901 tctctatata ctacgtatag gaaatgttta cattttcgta ttgttttcga ttcactctat
     3961 gaatagttct tactacaatt tttttgtcta aagagtaata ctagagataa acataaaaaa
     4021 tgtagaggtc gagtttagat gcaagttcaa ggagcgaaag gtggatgggt aggttatata
     4081 gggatatagc acagagatat atagcaaaga gatacttttg agcaatgttt gtggaagcgg
     4141 tattcgcaat attttagtag ctcgttacag tccggtgcgt ttttggtttt ttgaaagtgc
     4201 gtcttcagag cgcttttggt tttcaaaagc gctctgaagt tcctatactt tctagctaga
     4261 gaataggaac ttcggaatag gaacttcaaa gcgtttccga aaacgagcgc ttccgaaaat
     4321 gcaacgcgag ctgcgcacat acagctcact gttcacgtcg cacctatatc tgcgtgttgc
     4381 ctgtatatat atatacatga gaagaacggc atagtgcgtg tttatgctta aatgcgtact
     4441 tatatgcgtc tatttatgta ggatgaaagg tagtctagta cctcctgtga tattatccca
     4501 ttccatgcgg ggtatcgtat gcttccttca gcactaccct ttagctgttc tatatgctgc
     4561 cactcctcaa ttggattagt ctcatccttc aatgctatca tttcctttga tattggatcg
     4621 atccgatgat aagctgtcaa acatgagaat tgggtaataa ctgatataat taaattgaag
     4681 ctctaatttg tgagtttagt atacatgcat ttacttataa tacagttttt tagttttgct
     4741 ggccgcatct tctcaaatat gcttcccagc ctgcttttct gtaacgttca ccctctacct
     4801 tagcatccct tccctttgca aatagtcctc ttccaacaat aataatgtca gatcctgtag
     4861 agaccacatc atccacggtt ctatactgtt gacccaatgc gtctcccttg tcatctaaac
     4921 ccacaccggg tgtcataatc aaccaatcgt aaccttcatc tcttccaccc atgtctcttt
     4981 gagcaataaa gccgataaca aaatctttgt cgctcttcgc aatgtcaaca gtacccttag
     5041 tatattctcc agtagatagg gagcccttgc atgacaattc tgctaacatc aaaaggcctc
     5101 taggttcctt tgttacttct tctgccgcct gcttcaaacc gctaacaata cctgggccca
     5161 ccacaccgtg tgcattcgta atgtctgccc attctgctat tctgtataca cccgcagagt
     5221 actgcaattt gactgtatta ccaatgtcag caaattttct gtcttcgaag agtaaaaaat
     5281 tgtacttggc ggataatgcc tttagcggct taactgtgcc ctccatggaa aaatcagtca
     5341 agatatccac atgtgttttt agtaaacaaa ttttgggacc taatgcttca actaactcca
     5401 gtaattcctt ggtggtacga acatccaatg aagcacacaa gtttgtttgc ttttcgtgca
     5461 tgatattaaa tagcttggca gcaacaggac taggatgagt agcagcacgt tccttatatg
     5521 tagctttcga catgatttat cttcgtttcc tgcatgtttt tgttctgtgc agttgggtta
     5581 agaatactgg gcaatttcat gtttcttcaa cactacatat gcgtatatat accaatctaa
     5641 gtctgtgctc cttccttcgt tcttccttct gttcggagat taccgaatca aaaaaatttc
     5701 aaggaaaccg aaatcaaaaa aaagaataaa aaaaaaatga tgaattgaaa agctaattct
     5761 tgaagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     5821 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     5881 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     5941 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     6001 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     6061 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     6121 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     6181 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     6241 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     6301 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     6361 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     6421 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     6481 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     6541 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     6601 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     6661 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     6721 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     6781 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     6841 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     6901 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6961 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     7021 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     7081 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     7141 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     7201 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     7261 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     7321 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     7381 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     7441 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     7501 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     7561 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     7621 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     7681 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     7741 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
     7801 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     7861 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
     7921 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
     7981 tatgaccatg attacgccaa gcttaccgca tcaggaaatt gtaagcgtta atattttgtt
     8041 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     8101 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     8161 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     8221 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     8281 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     8341 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     8401 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     8461 acagggcgcg tccattcgc
//
