LOCUS       sequence-407703-        8437 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of ymTq2-P2A-ymScarletI in yeast.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8437)
  AUTHORS   Botman D, de Groot DH, Wehrens M, van Rossum K, Yska S, van der 
            Kieft B, Albert V, Minghetti N, Goedhart J, Teusink B
  TITLE     Side-by-side systematic characterization of novel FPs in budding 
            yeast
  JOURNAL   bioRxiv 2025.06.27.661975; doi: 
            https://doi.org/10.1101/2025.06.27.661975
REFERENCE   2  (bases 1 to 8437)
  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: "bioRxiv 
            2025.06.27.661975; doi: https://doi.org/10.1101/2025.06.27.661975"
COMMENT     Sequence Label: pDRF1-GW ymTq2-P2A-ymScarletI
FEATURES             Location/Qualifiers
     source          1..8437
                     /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             1557..1613
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     protein_bind    complement(2327..2351)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2413..2432)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2414..2432)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      2580..2767
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2635..2659)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(2779..2796)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2779..2795)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2788..2810)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3231..4577
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     CDS             complement(4688..5491)
                     /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(5492..5707)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(5732..5750)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5818..5922
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5923..5991,5992..6783)
                     /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(6141..6160)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6954..7542
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7443..7462
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7696..7713
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7830..7851
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7866..7871,7872..7889,7890..7896)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7904..7920
                     /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     7909..7931
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7928..7944
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7928..7944
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     rep_origin      complement(7974..8429)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8111..8132)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8323..8342
                     /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 atggttagta
      841 aaggtgaaga attgttcact ggtgtagtgc caattttagt agaattagat ggtgacgtta
      901 acggtcataa gttctctgta agcggtgaag gtgaaggtga cgcaacatac ggtaaattaa
      961 ctttgaaatt catttgtact acaggtaaat taccagttcc atggccaact ttggttacta
     1021 ctttgtcttg gggtgttcaa tgtttcgcta gatacccaga tcatatgaag caacatgatt
     1081 tctttaagtc tgctatgcca gaaggttacg ttcaagaaag aactatcttc ttcaaggatg
     1141 atggtaacta taagactaga gctgaagtta agttcgaagg agatacttta gttaatagaa
     1201 ttgaattgaa gggtattgat tttaaggaag atggtaatat tttgggtcat aagttggaat
     1261 ataattactt ctctgataac gtttacatta ctgctgataa gcaaaagaac ggtattaagg
     1321 cgaacttcaa gattagacat aacattgaag atggtggtgt tcaattagct gatcattacc
     1381 aacaaaacac tccaattggt gacggtccag ttttgttgcc agataaccat tacttgtcta
     1441 ctcaatctaa gttgtctaag gatccaaacg aaaagagaga tcatatggtt ttgttggaat
     1501 tcgttactgc tgctgggatc actttaggta tggatgaatt gtataaaggt tctggtgcaa
     1561 caaatttctc attgttgaag caggcgggag atgtggagga gaaccctgga cctcccggga
     1621 tggttagtaa aggtgaagct gttataaaag aattcatgag gtttaaagtt catatggaag
     1681 gttcaatgaa tggtcatgaa tttgaaattg aaggtgaagg tgaaggtaga ccatatgaag
     1741 gtacacaaac tgctaaattg aaagttacta aaggtggtcc attaccattt tcttgggata
     1801 ttttgtctcc acaattcatg tatggttcta gagcttttat taagcatcca gctgatattc
     1861 cagattatta taaacaatct tttcctgaag gttttaaatg ggaaagagtt atgaatttcg
     1921 aagatggtgg tgctgttact gttactcaag atacttcttt ggaagatggt actttaatct
     1981 ataaagttaa attgagaggt actaattttc caccagatgg tccagttatg caaaagaaaa
     2041 ctatgggttg ggaagcatct actgaaagat tgtatccaga agatggtgtt ttgaaaggag
     2101 acattaagat ggctttgaga ttgaaagatg gtggtcggta cctggccgac ttcaagacca
     2161 cctataaagc taaaaaacca gttcaaatgc caggtgcata taatgttgat agaaagttag
     2221 atataacgtc gcataacgag gactatactg tagttgaaca atatgaacgt agtgaaggta
     2281 gacatagtac cggaggaatg gatgaattgt ataaataagc ggccgcaccc agctttcttg
     2341 tacaaagtgg tgtagttaat tcatgatggc cgctgcaggt cgacctcgag ggggggcccg
     2401 gtacccaatt cgccctatag tgagtcgtat tacgcgcgga tccagctttg gacttcttcg
     2461 ccagaggttt ggtcaagtct ccaatcaagg ttgtcggctt gtctaccttg ccagaaattt
     2521 acgaaaagat ggaaaagggt caaatcgttg gtagatacgt tgttgacact tctaaataag
     2581 cgaatttctt atgatttatg atttttatta ttaaataagt tataaaaaaa ataagtgtat
     2641 acaaatttta aagtgactct taggttttaa aacgaaaatt cttattcttg agtaactctt
     2701 tcctgtaggt caggttgctt tctcaggtat agcatgaggt cgctcttatt gaccacacct
     2761 ctaccggcat gccaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     2821 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     2881 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgcct
     2941 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcataa tcggatcgta
     3001 cttgttaccc atcattgaat tttgaacatc cgaacctggg agttttccct gaaacagata
     3061 gtatatttga acctgtataa taatatatag tctagcgctt tacggaagac aatgtatgta
     3121 tttcggttcc tggagaaact attgcatcta ttgcataggt aatcttgcac gtcgcatccc
     3181 cggttcattt tctgcgtttc catcttgcac ttcaatagca tatctttgtt aacgaagcat
     3241 ctgtgcttca ttttgtagaa caaaaatgca acgcgagagc gctaattttt caaacaaaga
     3301 atctgagctg catttttaca gaacagaaat gcaacgcgaa agcgctattt taccaacgaa
     3361 gaatctgtgc ttcatttttg taaaacaaaa atgcaacgcg agagcgctaa tttttcaaac
     3421 aaagaatctg agctgcattt ttacagaaca gaaatgcaac gcgagagcgc tattttacca
     3481 acaaagaatc tatacttctt ttttgttcta caaaaatgca tcccgagagc gctatttttc
     3541 taacaaagca tcttagatta ctttttttct cctttgtgcg ctctataatg cagtctcttg
     3601 ataacttttt gcactgtagg tccgttaagg ttagaagaag gctactttgg tgtctatttt
     3661 ctcttccata aaaaaagcct gactccactt cccgcgttta ctgattacta gcgaagctgc
     3721 gggtgcattt tttcaagata aaggcatccc cgattatatt ctataccgat gtggattgcg
     3781 catactttgt gaacagaaag tgatagcgtt gatgattctt cattggtcag aaaattatga
     3841 acggtttctt ctattttgtc tctatatact acgtatagga aatgtttaca ttttcgtatt
     3901 gttttcgatt cactctatga atagttctta ctacaatttt tttgtctaaa gagtaatact
     3961 agagataaac ataaaaaatg tagaggtcga gtttagatgc aagttcaagg agcgaaaggt
     4021 ggatgggtag gttatatagg gatatagcac agagatatat agcaaagaga tacttttgag
     4081 caatgtttgt ggaagcggta ttcgcaatat tttagtagct cgttacagtc cggtgcgttt
     4141 ttggtttttt gaaagtgcgt cttcagagcg cttttggttt tcaaaagcgc tctgaagttc
     4201 ctatactttc tagctagaga ataggaactt cggaatagga acttcaaagc gtttccgaaa
     4261 acgagcgctt ccgaaaatgc aacgcgagct gcgcacatac agctcactgt tcacgtcgca
     4321 cctatatctg cgtgttgcct gtatatatat atacatgaga agaacggcat agtgcgtgtt
     4381 tatgcttaaa tgcgtactta tatgcgtcta tttatgtagg atgaaaggta gtctagtacc
     4441 tcctgtgata ttatcccatt ccatgcgggg tatcgtatgc ttccttcagc actacccttt
     4501 agctgttcta tatgctgcca ctcctcaatt ggattagtct catccttcaa tgctatcatt
     4561 tcctttgata ttggatcgat ccgatgataa gctgtcaaac atgagaattg ggtaataact
     4621 gatataatta aattgaagct ctaatttgtg agtttagtat acatgcattt acttataata
     4681 cagtttttta gttttgctgg ccgcatcttc tcaaatatgc ttcccagcct gcttttctgt
     4741 aacgttcacc ctctacctta gcatcccttc cctttgcaaa tagtcctctt ccaacaataa
     4801 taatgtcaga tcctgtagag accacatcat ccacggttct atactgttga cccaatgcgt
     4861 ctcccttgtc atctaaaccc acaccgggtg tcataatcaa ccaatcgtaa ccttcatctc
     4921 ttccacccat gtctctttga gcaataaagc cgataacaaa atctttgtcg ctcttcgcaa
     4981 tgtcaacagt acccttagta tattctccag tagataggga gcccttgcat gacaattctg
     5041 ctaacatcaa aaggcctcta ggttcctttg ttacttcttc tgccgcctgc ttcaaaccgc
     5101 taacaatacc tgggcccacc acaccgtgtg cattcgtaat gtctgcccat tctgctattc
     5161 tgtatacacc cgcagagtac tgcaatttga ctgtattacc aatgtcagca aattttctgt
     5221 cttcgaagag taaaaaattg tacttggcgg ataatgcctt tagcggctta actgtgccct
     5281 ccatggaaaa atcagtcaag atatccacat gtgtttttag taaacaaatt ttgggaccta
     5341 atgcttcaac taactccagt aattccttgg tggtacgaac atccaatgaa gcacacaagt
     5401 ttgtttgctt ttcgtgcatg atattaaata gcttggcagc aacaggacta ggatgagtag
     5461 cagcacgttc cttatatgta gctttcgaca tgatttatct tcgtttcctg catgtttttg
     5521 ttctgtgcag ttgggttaag aatactgggc aatttcatgt ttcttcaaca ctacatatgc
     5581 gtatatatac caatctaagt ctgtgctcct tccttcgttc ttccttctgt tcggagatta
     5641 ccgaatcaaa aaaatttcaa ggaaaccgaa atcaaaaaaa agaataaaaa aaaaatgatg
     5701 aattgaaaag ctaattcttg aagacgaaag ggcctcgtga tacgcctatt tttataggtt
     5761 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     5821 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     5881 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     5941 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     6001 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     6061 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     6121 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     6181 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     6241 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     6301 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     6361 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     6421 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     6481 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     6541 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     6601 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     6661 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     6721 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     6781 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     6841 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     6901 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     6961 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     7021 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     7081 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     7141 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     7201 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     7261 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     7321 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     7381 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     7441 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     7501 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     7561 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     7621 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     7681 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     7741 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     7801 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     7861 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     7921 tttcacacag gaaacagcta tgaccatgat tacgccaagc ttaccgcatc aggaaattgt
     7981 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     8041 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     8101 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     8161 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     8221 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     8281 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     8341 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     8401 cgcgcttaat gcgccgctac agggcgcgtc cattcgc
//
