LOCUS       sequence-223804-        8467 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Produces equimolar expression of yeast codon-optimized mScarletI and
            mTq2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8467)
  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 8467)
  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 ymScarletI-T2A-mTurquoise2
FEATURES             Location/Qualifiers
     source          1..8467
                     /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             1553..1606
                     /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      1622..1631
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1628..1630,1631..1633,1634..2347)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2012)"
                     /label=mTurquoise2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1673..1694)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1934..1953)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2281..2302
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(2357..2381)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2443..2462)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2444..2462)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      2610..2797
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2665..2689)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(2809..2826)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2809..2825)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2818..2840)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3261..4607
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     CDS             complement(4718..5521)
                     /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(5522..5737)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(5762..5780)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5848..5952
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5953..6021,6022..6813)
                     /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(6171..6190)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6984..7572
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7473..7492
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7726..7743
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7860..7881
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7896..7901,7902..7919,7920..7926)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7934..7950
                     /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     7939..7961
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7958..7974
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7958..7974
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     rep_origin      complement(8004..8459)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8141..8162)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8353..8372
                     /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 aaggtgaagc tgttataaaa gaattcatga ggtttaaagt
      901 tcatatggaa ggttcaatga atggtcatga atttgaaatt gaaggtgaag gtgaaggtag
      961 accatatgaa ggtacacaaa ctgctaaatt gaaagttact aaaggtggtc cattaccatt
     1021 ttcttgggat attttgtctc cacaattcat gtatggttct agagctttta ttaagcatcc
     1081 agctgatatt ccagattatt ataaacaatc ttttcctgaa ggttttaaat gggaaagagt
     1141 tatgaatttc gaagatggtg gtgctgttac tgttactcaa gatacttctt tggaagatgg
     1201 tactttaatc tataaagtta aattgagagg tactaatttt ccaccagatg gtccagttat
     1261 gcaaaagaaa actatgggtt gggaagcatc tactgaaaga ttgtatccag aagatggtgt
     1321 tttgaaagga gacattaaga tggctttgag attgaaagat ggtggtcggt acctggccga
     1381 cttcaagacc acctataaag ctaaaaaacc agttcaaatg ccaggtgcat ataatgttga
     1441 tagaaagtta gatataacgt cgcataacga ggactatact gtagttgaac aatatgaacg
     1501 tagtgaaggt agacatagta ccggaggaat ggatgaattg tataaatccg gagagggcag
     1561 aggaagtctt ctaacatgcg gtgacgtgga ggagaatccc ggccctgggg atccaccggt
     1621 cgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     1681 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     1741 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     1801 gcccaccctc gtgaccaccc tgtcctgggg cgtgcagtgc ttcgcccgct accccgacca
     1861 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     1921 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     1981 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     2041 ggggcacaag ctggagtaca actactttag cgacaacgtc tatatcaccg ccgacaagca
     2101 gaagaacggc atcaaggcca acttcaagat ccgccacaac atcgaggacg gcggcgtgca
     2161 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     2221 caaccactac ctgagcaccc agtccaagct gagcaaagac cccaacgaga agcgcgatca
     2281 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     2341 caagtaaagc ggccgcaccc agctttcttg tacaaagtgg tgtagttaat tcatgatggc
     2401 cgctgcaggt cgacctcgag ggggggcccg gtacccaatt cgccctatag tgagtcgtat
     2461 tacgcgcgga tccagctttg gacttcttcg ccagaggttt ggtcaagtct ccaatcaagg
     2521 ttgtcggctt gtctaccttg ccagaaattt acgaaaagat ggaaaagggt caaatcgttg
     2581 gtagatacgt tgttgacact tctaaataag cgaatttctt atgatttatg atttttatta
     2641 ttaaataagt tataaaaaaa ataagtgtat acaaatttta aagtgactct taggttttaa
     2701 aacganaatt cttattcttg agtaactctt tcctgtaggt caggttgctt tctcaggtat
     2761 agcatgaggt cgctcttatt gaccacacct ctaccggcat gccaattcac tggccgtcgt
     2821 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     2881 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     2941 gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg
     3001 cggtatttca caccgcataa tcggatcgta cttgttaccc atcattgaat tttgaacatc
     3061 cgaacctggg agttttccct gaaacagata gtatatttga acctgtataa taatatatag
     3121 tctagcgctt tacggaagac aatgtatgta tttcggttcc tggagaaact attgcatcta
     3181 ttgcataggt aatcttgcac gtcgcatccc cggttcattt tctgcgtttc catcttgcac
     3241 ttcaatagca tatctttgtt aacgaagcat ctgtgcttca ttttgtagaa caaaaatgca
     3301 acgcgagagc gctaattttt caaacaaaga atctgagctg catttttaca gaacagaaat
     3361 gcaacgcgaa agcgctattt taccaacgaa gaatctgtgc ttcatttttg taaaacaaaa
     3421 atgcaacgcg agagcgctaa tttttcaaac aaagaatctg agctgcattt ttacagaaca
     3481 gaaatgcaac gcgagagcgc tattttacca acaaagaatc tatacttctt ttttgttcta
     3541 caaaaatgca tcccgagagc gctatttttc taacaaagca tcttagatta ctttttttct
     3601 cctttgtgcg ctctataatg cagtctcttg ataacttttt gcactgtagg tccgttaagg
     3661 ttagaagaag gctactttgg tgtctatttt ctcttccata aaaaaagcct gactccactt
     3721 cccgcgttta ctgattacta gcgaagctgc gggtgcattt tttcaagata aaggcatccc
     3781 cgattatatt ctataccgat gtggattgcg catactttgt gaacagaaag tgatagcgtt
     3841 gatgattctt cattggtcag aaaattatga acggtttctt ctattttgtc tctatatact
     3901 acgtatagga aatgtttaca ttttcgtatt gttttcgatt cactctatga atagttctta
     3961 ctacaatttt tttgtctaaa gagtaatact agagataaac ataaaaaatg tagaggtcga
     4021 gtttagatgc aagttcaagg agcgaaaggt ggatgggtag gttatatagg gatatagcac
     4081 agagatatat agcaaagaga tacttttgag caatgtttgt ggaagcggta ttcgcaatat
     4141 tttagtagct cgttacagtc cggtgcgttt ttggtttttt gaaagtgcgt cttcagagcg
     4201 cttttggttt tcaaaagcgc tctgaagttc ctatactttc tagctagaga ataggaactt
     4261 cggaatagga acttcaaagc gtttccgaaa acgagcgctt ccgaaaatgc aacgcgagct
     4321 gcgcacatac agctcactgt tcacgtcgca cctatatctg cgtgttgcct gtatatatat
     4381 atacatgaga agaacggcat agtgcgtgtt tatgcttaaa tgcgtactta tatgcgtcta
     4441 tttatgtagg atgaaaggta gtctagtacc tcctgtgata ttatcccatt ccatgcgggg
     4501 tatcgtatgc ttccttcagc actacccttt agctgttcta tatgctgcca ctcctcaatt
     4561 ggattagtct catccttcaa tgctatcatt tcctttgata ttggatcgat ccgatgataa
     4621 gctgtcaaac atgagaattg ggtaataact gatataatta aattgaagct ctaatttgtg
     4681 agtttagtat acatgcattt acttataata cagtttttta gttttgctgg ccgcatcttc
     4741 tcaaatatgc ttcccagcct gcttttctgt aacgttcacc ctctacctta gcatcccttc
     4801 cctttgcaaa tagtcctctt ccaacaataa taatgtcaga tcctgtagag accacatcat
     4861 ccacggttct atactgttga cccaatgcgt ctcccttgtc atctaaaccc acaccgggtg
     4921 tcataatcaa ccaatcgtaa ccttcatctc ttccacccat gtctctttga gcaataaagc
     4981 cgataacaaa atctttgtcg ctcttcgcaa tgtcaacagt acccttagta tattctccag
     5041 tagataggga gcccttgcat gacaattctg ctaacatcaa aaggcctcta ggttcctttg
     5101 ttacttcttc tgccgcctgc ttcaaaccgc taacaatacc tgggcccacc acaccgtgtg
     5161 cattcgtaat gtctgcccat tctgctattc tgtatacacc cgcagagtac tgcaatttga
     5221 ctgtattacc aatgtcagca aattttctgt cttcgaagag taaaaaattg tacttggcgg
     5281 ataatgcctt tagcggctta actgtgccct ccatggaaaa atcagtcaag atatccacat
     5341 gtgtttttag taaacaaatt ttgggaccta atgcttcaac taactccagt aattccttgg
     5401 tggtacgaac atccaatgaa gcacacaagt ttgtttgctt ttcgtgcatg atattaaata
     5461 gcttggcagc aacaggacta ggatgagtag cagcacgttc cttatatgta gctttcgaca
     5521 tgatttatct tcgtttcctg catgtttttg ttctgtgcag ttgggttaag aatactgggc
     5581 aatttcatgt ttcttcaaca ctacatatgc gtatatatac caatctaagt ctgtgctcct
     5641 tccttcgttc ttccttctgt tcggagatta ccgaatcaaa aaaatttcaa ggaaaccgaa
     5701 atcaaaaaaa agaataaaaa aaaaatgatg aattgaaaag ctaattcttg aagacgaaag
     5761 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     5821 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     5881 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5941 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     6001 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     6061 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     6121 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     6181 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     6241 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     6301 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     6361 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     6421 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6481 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6541 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6601 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6661 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6721 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6781 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6841 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     6901 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     6961 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     7021 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     7081 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     7141 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     7201 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     7261 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     7321 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     7381 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     7441 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     7501 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     7561 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     7621 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     7681 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     7741 gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat
     7801 taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt
     7861 aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt
     7921 atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat
     7981 tacgccaagc ttaccgcatc aggaaattgt aagcgttaat attttgttaa aattcgcgtt
     8041 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     8101 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     8161 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     8221 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     8281 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     8341 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     8401 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     8461 cattcgc
//
