LOCUS       sequence-244230-        8218 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A dual reporter vector modified from Sharon et al (Nature Biotech, 
            2012) containing Ura3 and Nat1 selectable markers, a constant 
            background RFP (TEF2::mCherry), and a variable YFP 
            (pTpA+MCS::yEVenus)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8218)
  AUTHORS   de Boer CG, Vaishnav ED, Sadeh R, Abeyta EL, Friedman N, Regev A
  TITLE     Deciphering eukaryotic gene-regulatory logic with 100 million random
            promoters.
  JOURNAL   Nat Biotechnol. 2020 Jan;38(1):56-65. doi: 
            10.1038/s41587-019-0315-8. Epub 2019 Dec 2.
  PUBMED    31792407
REFERENCE   2  (bases 1 to 8218)
  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: 
            "10.1038/s41587-019-0315-8"; journalName: "Nat Biotechnol"; date: 
            "2020-01"; volume: "38"; issue: "1"; pages: "56-65"
COMMENT     Sequence Label: yeast_DualReporter
FEATURES             Location/Qualifiers
     source          1..8218
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(103..894,895..963))
                     /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     726..745
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(964..1068)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(1133..1320)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     CDS             complement(1347..2057)
                     /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(1625..1644)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1894..1912
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(2674..2696)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        2896..2914
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2896..2913
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     promoter        2988..3203
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             3204..4007
                     /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"
     primer_bind     complement(4275..4296)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             4541..5257
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      5278..5465
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     gene            5540..6659
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        5540..5883
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5884..6456
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      6462..6659
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     misc_feature    6726..7229
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     complement(7285..7304)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7286..7304)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7391..7408)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7562..8150)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7642..7661)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
       61 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
      121 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
      181 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
      241 gcgagaccca cgttcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
      301 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
      361 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
      421 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
      481 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
      541 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
      601 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
      661 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
      721 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
      781 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
      841 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
      901 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
      961 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     1021 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     1081 aaaagtgcca cctgacgtca tctatattac cctgttatcc ctagcggatc tgccggtaga
     1141 ggtgtggtca ataagagcga cctcatacta tacctgagaa agcaacctga cctacaggaa
     1201 agagttactc aagaataaga attttcgttt taaaacctaa gagtcacttt aaaatttgta
     1261 tacacttatt ttttttataa cttatttaat aataaaaatc ataaatcata agaaattcgc
     1321 ttatttagaa gtggcgcgcc ggtccgttac ttgtacagct cgtccatgcc gccggtggag
     1381 tggcggccct cggcgcgttc gtactgttcc acgatggtgt agtcctcgtt gtgggaggtg
     1441 atgtccaact tgatgttgac gttgtaggcg ccgggcagct gcacgggctt cttggccttg
     1501 taggtggtct tgacctcagc gtcgtagtgg ccgccgtcct tcagcttcag cctctgcttg
     1561 atctcgccct tcagggcgcc gtcctcgggg tacatccgct cggaggaggc ctcccagccc
     1621 atggtcttct tctgcattac ggggccgtcg gaggggaagt tggtgccgcg cagcttcacc
     1681 ttgtagatga actcgccgtc ctgcagggag gagtcctggg tcacggtcac cacgccgccg
     1741 tcctcgaagt tcatcacgcg ctcccacttg aagccctcgg ggaaggacag cttcaagtag
     1801 tcggggatgt cggcggggtg cttcacgtag gccttggagc cgtacatgaa ctgaggggac
     1861 aggatgtccc aggcgaaggg cagggggcca cccttggtca ccttcagctt ggcggtctgg
     1921 gtgccctcgt aggggcggcc ctcgccctcg ccctcgatct cgaactcgtg gccgttcacg
     1981 gagccctcca tgtgcacctt gaagcgcatg aactccttga tgatggccat gttatcctcc
     2041 tcgcccttgc tcaccatggt actagtgttt agttaattat agttcgttga ccgtatattc
     2101 taaaaacaag tactccttaa aaaaaaacct tgaagggaat aaacaagtag aatagataga
     2161 gagaaaaata gaaaatgcaa gagaatttat atattagaaa gagagaaaga aaaatggaaa
     2221 aaaaaaaata ggaaaagcca gaaatagcac tagaaggagc gacaccagaa aagaaggtga
     2281 tggaaccaat ttagctatat atagttaact accggctcga tcatctctgc ctccagcata
     2341 gtcgaagaag aatttttttt ttcttgaggc ttctgtcagc aactcgtatt ttttctttct
     2401 tttttggtga gcctaaaaag ttcccacgtt ctcttgtacg acgccgtcac aaacaacctt
     2461 atgggtaatt tgtcgcggtc tgggtgtata aatgtgtggg tgcaacatga atgtacggag
     2521 gtagtttgct gattggcggt ctatagatac cttggttatg gcgccctcac agccggcagg
     2581 ggaagcgcct acgcttgaca tctactatat gtaagtatac ggccccatat ataggccctt
     2641 tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
     2701 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
     2761 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
     2821 agtgcaccat atggacatat tgtcgttaga acgcggctac aattaataca taaccttatg
     2881 tatcatacac atacgattta ggtgacacta tagaacgcgg ccgccagctg aagctttaac
     2941 tatgcggcat cagagcagat tgtactgaga gtgcaccata ccaccttttc aattcatcat
     3001 ttttttttta ttcttttttt tgatttcggt ttccttgaaa tttttttgat tcggtaatct
     3061 ccgaacagaa ggaagaacga aggaaggagc acagacttag attggtatat atacgcatat
     3121 gtagtgttga agaaacatga aattgcccag tattcttaac ccaactgcac agaacaaaaa
     3181 cctgcaggaa acgaagataa atcatgtcga aagctacata taaggaacgt gctgctactc
     3241 atcctagtcc tgttgctgcc aagctattta atatcatgca cgaaaagcaa acaaacttgt
     3301 gtgcttcatt ggatgttcgt accaccaagg aattactgga gttagttgaa gcattaggtc
     3361 ccaaaatttg tttactaaaa acacatgtgg atatcttgac tgatttttcc atggagggca
     3421 cagttaagcc gctaaaggca ttatccgcca agtacaattt tttactcttc gaagacagaa
     3481 aatttgctga cattggtaat acagtcaaat tgcagtactc tgcgggtgta tacagaatag
     3541 cagaatgggc agacattacg aatgcacacg gtgtggtggg cccaggtatt gttagcggtt
     3601 tgaagcaggc ggcagaagaa gtaacaaagg aacctagagg ccttttgatg ttagcagaat
     3661 tgtcatgcaa gggctcccta tctactggag aatatactaa gggtactgtt gacattgcga
     3721 agagcgacaa agattttgtt atcggcttta ttgctcaaag agacatgggt ggaagagatg
     3781 aaggttacga ttggttgatt atgacacccg gtgtgggttt agatgacaag ggagacgcat
     3841 tgggtcaaca gtatagaacc gtggatgatg tggtctctac aggatctgac attattattg
     3901 ttggaagagg actatttgca aagggaaggg atgctaaggt agagggtgaa cgttacagaa
     3961 aagcaggctg ggaagcatat ttgagaagat gcggccagca aaactaaaaa actgtattat
     4021 aagtaaatgc atgtatacta aactcacaaa ttagagcttc aatttaatta tatcagttat
     4081 taccctatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggaaa
     4141 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     4201 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     4261 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     4321 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     4381 caagtgctag caggaatgat gcaaaaggtt cccgattcga actgcatttt tttcacatct
     4441 cgaggttacg gctgtttctt aattaaaaaa agatagaaaa cattaggagt gtaacacaag
     4501 actttcggat cctgagcagg caagataaac gaaggcaaag atgtctaaag gtgaagaatt
     4561 attcactggt gttgtcccaa ttttggttga attagatggt gatgttaatg gtcacaaatt
     4621 ttctgtctcc ggtgaaggtg aaggtgatgc tacttacggt aaattgacct taaaattgat
     4681 ttgtactact ggtaaattgc cagttccatg gccaacctta gtcactactt taggttatgg
     4741 tttgcaatgt tttgctagat acccagatca tatgaaacaa catgactttt tcaagtctgc
     4801 catgccagaa ggttatgttc aagaaagaac tatttttttc aaagatgacg gtaactacaa
     4861 gaccagagct gaagtcaagt ttgaaggtga taccttagtt aatagaatcg aattaaaagg
     4921 tattgatttt aaagaagatg gtaacatttt aggtcacaaa ttggaataca actataactc
     4981 tcacaatgtt tacatcactg ctgacaaaca aaagaatggt atcaaagcta acttcaaaat
     5041 tagacacaac attgaagatg gtggtgttca attagctgac cattatcaac aaaatactcc
     5101 aattggtgat ggtccagtct tgttaccaga caaccattac ttatcctatc aatctgcctt
     5161 atccaaagat ccaaacgaaa agagagacca catggtcttg ttagaatttg ttactgctgc
     5221 tggtattacc catggtatgg atgaattgta caaataaggc gcgccacttc taaataagcg
     5281 aatttcttat gatttatgat ttttattatt aaataagtta taaaaaaaat aagtgtatac
     5341 aaattttaaa gtgactctta ggttttaaaa cgaaaattct tattcttgag taactctttc
     5401 ctgtaggtca ggttgctttc tcaggtatag tatgaggtcg ctcttattga ccacacctct
     5461 accggcagat ccgctaggga taacagggta atatagatct gtttagcttg cctcgtcccc
     5521 gccgggtcac ccggccagcg acatggaggc ccagaatacc ctccttgaca gtcttgacgt
     5581 gcgcagctca ggggcatgat gtgactgtcg cccgtacatt tagcccatac atccccatgt
     5641 ataatcattt gcatccatac attttgatgg ccgcacggcg cgaagcaaaa attacggctc
     5701 ctcgctgcag acctgcgagc agggaaacgc tcccctcaca gacgcgttga attgtcccca
     5761 cgccgcgccc ctgtagagaa atataaaagg ttaggatttg ccactgaggt tcttctttca
     5821 tatacttcct tttaaaatct tgctaggata cagttctcac atcacatccg aacataaaca
     5881 accatgggta ccactcttga cgacacggct taccggtacc gcaccagtgt cccgggggac
     5941 gccgaggcca tcgaggcact ggatgggtcc ttcaccaccg acaccgtctt ccgcgtcacc
     6001 gccaccgggg acggcttcac cctgcgggag gtgccggtgg acccgcccct gaccaaggtg
     6061 ttccccgacg acgaatcgga cgacgaatcg gacgacgggg aggacggcga cccggactcc
     6121 cggacgttcg tcgcgtacgg ggacgacggc gacctggcgg gcttcgtggt cgtctcgtac
     6181 tccggctgga accgccggct gaccgtcgag gacatcgagg tcgccccgga gcaccggggg
     6241 cacggggtcg ggcgcgcgtt gatggggctc gcgacggagt tcgcccgcga gcggggcgcc
     6301 gggcacctct ggctggaggt caccaacgtc aacgcaccgg cgatccacgc gtaccggcgg
     6361 atggggttca ccctctgcgg cctggacacc gccctgtacg acggcaccgc ctcggacggc
     6421 gagcaggcgc tctacatgag catgccctgc ccctaatcag tactgacaat aaaaagattc
     6481 ttgttttcaa gaacttgtca tttgtatagt ttttttatat tgtagttgtt ctattttaat
     6541 caaatgttag cgtgatttat attttttttc gcctcgacat catctgccca gatgcgaagt
     6601 taagtgcgca gaaagtaata tcatgcgtca atcgtatgtg aatgctggtc gctatactgc
     6661 tgtcgattcg atactaacgc cgccatccag tgtcgaaaac gagctcgaat tcctgggtcc
     6721 ttttcatcac gtgctataaa aataattata atttaaattt tttaatataa atatataaat
     6781 taaaaataga aagtaaaaaa agaaattaaa gaaaaaatag tttttgtttt ccgaagatgt
     6841 aaaagactct agggggatcg ccaacaaata ctacctttta tcttgctctt cctgctctca
     6901 ggtattaatg ccgaattgtt tcatcttgtc tgtgtagaag accacacacg aaaatcctgt
     6961 gattttacat tttacttatc gttaatcgaa tgtatatcta tttaatctgc ttttcttgtc
     7021 taataaatat atatgtaaag tacgcttttt gttgaaattt tttaaacctt tgtttatttt
     7081 tttttcttca ttccgtaact cttctacctt ctttatttac tttctaaaat ccaaatacaa
     7141 aacataaaaa taaataaaca cagagtaaat tcccaaatta ttccatcatt aaaagatacg
     7201 aggcgcgtgt aagttacagg caagcgatcc gtccgatatc atcagatcca ctagtggcct
     7261 atgcggccgc ggatctgccg gtctccctat agtgagtcgt attaatttcg ataagccagg
     7321 ttaacctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     7381 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     7441 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     7501 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     7561 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     7621 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     7681 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     7741 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     7801 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     7861 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     7921 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     7981 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     8041 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     8101 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     8161 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggatttt
//
