LOCUS       sequence-360583-        7214 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeat two-hybrid fluorecent reporter for genomic integration.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7214)
  AUTHORS   Feuer E, Zimran G, Shpilman M, Mosquna A
  TITLE     A Modified Yeast Two-Hybrid Platform Enables Dynamic Control of 
            Expression Intensities to Unmask Properties of Protein-Protein 
            Interactions.
  JOURNAL   ACS Synth Biol. 2022 Jul 27. doi: 10.1021/acssynbio.2c00192.
  PUBMED    35895499
REFERENCE   2  (bases 1 to 7214)
  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: "ACS Synth 
            Biol. 2022 Jul 27. doi: 10.1021/acssynbio.2c00192."
COMMENT     Sequence Label: pHO-UAS-mScarlet-I
FEATURES             Location/Qualifiers
     source          1..7214
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             336..1145
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(403..422)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1151..1348
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     1918..1934
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1918..1934
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(1948..1967)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1949..1967)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1974..1991)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1974..1990)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1983..2005)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2290..2309)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2422..2443
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2597..2616)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2716..2738
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2776..2794)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2862..2966
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2967..3035,3036..3827)
                     /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(3185..3204)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3998..4586
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4487..4506
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4740..4757
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4874..4895
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    4950..4966
                     /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     4955..4977
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4974..4990
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4974..4990
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    6049..6166
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     CDS             6292..6987
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(6425..6443)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     6693..6712
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     terminator      7006..7193
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(7061..7085)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            join(7206..7214,1..1348)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        join(7206..7214,1..335)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
ORIGIN
        1 gcccagaata ccctccttga cagtcttgac gtgcgcagct caggggcatg atgtgactgt
       61 cgcccgtaca tttagcccat acatccccat gtataatcat ttgcatccat acattttgat
      121 ggccgcacgg cgcgaagcaa aaattacggc tcctcgctgc agacctgcga gcagggaaac
      181 gctcccctca cagacgcgtt gaattgtccc cacgccgcgc ccctgtagag aaatataaaa
      241 ggttaggatt tgccactgag gttcttcttt catatacttc cttttaaaat cttgctagga
      301 tacagttctc acatcacatc cgaacataaa caaccatggg taaggaaaag actcacgttt
      361 cgaggccgcg attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg
      421 ataatgtcgg gcaatcaggt gcgacaatct atcgattgta tgggaagccc gatgcgccag
      481 agttgtttct gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca
      541 gactaaactg gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc
      601 ctgatgatgc atggttactc accactgcga tccccggcaa aacagcattc caggtattag
      661 aagaatatcc tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt
      721 tgcattcgat tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc
      781 aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta
      841 atggctggcc tgttgaacaa gtctggaaag aaatgcataa gcttttgcca ttctcaccgg
      901 attcagtcgt cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat
      961 taataggttg tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca
     1021 tcctatggaa ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat
     1081 atggtattga taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt
     1141 tctaatcagt actgacaata aaaagattct tgttttcaag aacttgtcat ttgtatagtt
     1201 tttttatatt gtagttgttc tattttaatc aaatgttagc gtgatttata ttttttttcg
     1261 cctcgacatc atctgcccag atgcgaagtt aagtgcgcag aaagtaatat catgcgtcaa
     1321 tcgtatgtga atgctggtcg ctatactgct gtcgattcga tactaacgcc gccatccagt
     1381 gtcgaaaacg agctcgaatt cctgggggaa caacttcaca gaatgttttg tcatattgtc
     1441 gaagtggtca caaaacaaga gaagttccgc caattataaa aagggaaccc gtatatttca
     1501 gcttcacgga tgatttccag ggtgagagta ctgtatatgg gcttacgata gaaggccata
     1561 aaaatttctt gcttggcaac aaaatagaag tgaaatcatg tcgaggctgc tgtgtgggag
     1621 aacagcataa aatatcacaa aaaaagaatc taaaacactg tgttgcttgt cccagaaagg
     1681 gaatcaagta tttttataaa gattggagtg gtaaaaatcg agtatgtgct agatgctatg
     1741 gaagatacaa attcagcggt catcactgta taaattgcaa gtatgtacca gaagcacgtg
     1801 aagtgaaaaa ggcaaaagac aaaggcgaaa aattgggcat tacgcccgaa ggtttgccag
     1861 ttaaaggacc agagtgtata aaatgtggcg gaatcttaca gtggcctatg cggccgctct
     1921 agaactagtg gatcgatccc caattcgccc tatagtgagt cgtattacaa ttcactggcc
     1981 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca
     2041 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc
     2101 caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat
     2161 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     2221 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct
     2281 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     2341 gtcaagctct aaatcggggt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     2401 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta
     2461 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa
     2521 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     2581 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     2641 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     2701 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     2761 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     2821 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     2881 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     2941 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     3001 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3061 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3121 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3181 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     3241 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3301 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3361 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3421 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3481 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     3541 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     3601 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     3661 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     3721 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     3781 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     3841 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     3901 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     3961 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4021 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4081 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4141 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4201 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4261 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4321 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4381 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4441 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4501 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4561 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     4621 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     4681 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     4741 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc
     4801 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     4861 agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta
     4921 tgcttccgcg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
     4981 cagctatgac catgattacg ccactagtcc gaggcctcga gatccgatat cgccgtggcg
     5041 gccgccagct gaagcttaat tatcctgggc acgagtgaaa caaagctaaa acctttattt
     5101 agcatggcca ttgaatgtaa caattatata tatcgcaagc acaaaaaatc aaggagagag
     5161 aactaccact ttgttcatgt gtacaatgtt cattatctcc ataagcaaaa aaaaaaaaat
     5221 agaaaacata tgctataagg ttgatattct cacgagtaag cggcacttgc tacttattga
     5281 cattgcagat ttttggctac agaaatagta tattagagat tataattgct aatcaaatca
     5341 aaatataaaa ttagtaaacc aaaccattta tacccttcct tagtagttat ggattgtttt
     5401 ttaatgatat ttctgcaaac caaagaaaga ttgttatcca gatagaattt agttttgata
     5461 ttcatttttt tgttgaagat tgaacgccat atctgggcct cataattcaa aagacggtgc
     5521 cattatcggt agcgtttcgc attgtactgg atttcagaaa tttcacagtt gatgaatcga
     5581 aaagaatggt ctcattgcaa cacgtaaggt taagatgtcc ctttttacca ttataggcaa
     5641 taaatgaatc ataaaacgac cgtatactgg tgaaatagta gggagaacga gtacctgtag
     5701 taaaaagtat aaatcatagt taatcgggca atgtccctcg atcaaggagt attgtgtcat
     5761 gttcgagaca aacgccaaca tttttgtttc ttttggacaa atgttgtttg catttatgat
     5821 ccgttatatt ttgatctaat gtagagttgc acgtagttct tactggcaaa gaaatcgatg
     5881 cataccaaaa aagaataaag gtgatatttg atctttaccg tttagttcca acgtaaaatt
     5941 gtgcctttgg acttaaaatg gcgtcgtacg ctgcaggtcg acgcttccaa gcaggttgag
     6001 acaaatcgga tccggggaat tccctagtat tgtagaatct ttattgttcg gagcagtgcg
     6061 gcgcgaggca catctgcgtt tcaggaacgc gaccggtgaa gacgaggacg cacggaggag
     6121 agtcttcctt cggagggctg tcacccgctc ggcggcttct aatccgtgga attccttttt
     6181 ttttcttagc gattggcatt atcacataat gaattataca ttatataaag taatgtgatt
     6241 tcttcgaaga atatactaaa aaatgagcag gcaagataaa cgaaggcaaa gatggtgagc
     6301 aagggcgagg cagtgatcaa ggagttcatg cggttcaagg tgcacatgga gggctccatg
     6361 aacggccacg agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag
     6421 accgccaagc tgaaggtgac caagggtggc cccctgccct tctcctggga catcctgtcc
     6481 cctcagttca tgtacggctc cagggccttc atcaagcacc ccgccgacat ccccgactac
     6541 tataagcagt ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc
     6601 ggcgccgtga ccgtgaccca ggacacctcc ctggaggacg gcaccctgat ctacaaggtg
     6661 aagctccgcg gcaccaactt ccctcctgac ggccccgtaa tgcagaagaa gacaatgggc
     6721 tgggaagcgt ccaccgagcg gttgtacccc gaggacggcg tgctgaaggg cgacattaag
     6781 atggccctgc gcctgaagga cggcggccgt tacctggcgg acttcaagac cacctacaag
     6841 gccaagaagc ccgtgcagat gcccggcgcc tacaacgtgg accgcaagtt ggacatcacc
     6901 tcccacaacg aggactacac cgtggtggaa cagtacgaac gctccgaggg ccgccactcc
     6961 accggcggca tggacgagct gtacaagtag ccaagctaat tccgggcgaa tttcttatga
     7021 tttatgattt ttattattaa ataagttata aaaaaaataa gtgtatacaa attttaaagt
     7081 gactcttagg ttttaaaacg aaaattctta ttcttgagta actctttcct gtaggtcagg
     7141 ttgctttctc aggtatagca tgaggtcgct cttattgacc acacctctac cggcatccgg
     7201 ccagcgacat ggag
//
