LOCUS       sequence-259392-        8787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes VP16-CIB1 under pADH1 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8787)
  AUTHORS   An-Adirekkun JM, Stewart CJ, Geller SH, Patel MT, Melendez J, Oakes 
            BL, Noyes MB, McClean MN
  TITLE     A yeast optogenetic toolkit (yOTK) for gene expression control in 
            Saccharomyces cerevisiae.
  JOURNAL   Biotechnol Bioeng. 2019 Dec 2. doi: 10.1002/bit.27234.
  PUBMED    31788779
REFERENCE   2  (bases 1 to 8787)
  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: "Biotechnol 
            Bioeng. 2019 Dec 2. doi: 10.1002/bit.27234."
COMMENT     Sequence Label: pMM0281
FEATURES             Location/Qualifiers
     source          1..8787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1476..2180
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             2226..2246
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     2474..2493
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2474..2492
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2517..3026
                     /codon_start=1
                     /product="N-terminal portion of the CIB1 transcription 
                     factor from Arabidopsis thaliana"
                     /label=CIBN
                     /note="binds to CRY2 (cryptochrome 2) that has absorbed 
                     blue light (Kennedy et al., 2010)"
                     /translation="MNGAIGGDLLLNFPDMSVLERQRAHLKYLNPTFDSPLAGFFADSS
                     MITGGEMDSYLSTAGLNLPMMYGETTVEGDSRLSISPETTLGTGNFKKRKFDTETKDCN
                     EKKKKMTMNRDDLVEEGEEEKSKITEQNNGSTKSIKKMKHKAKKEENNFSNDSSKVTKE
                     LEKTDYI"
     terminator      3920..4107
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(3975..3999)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(4224..5318)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIEATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(5319..5724)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(5766..5782)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5766..5782)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5779..5801)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5790..5806
                     /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)."
     promoter        complement(join(5814..5820,5821..5838,5839..5844))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5859..5880
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    5935..5968
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    6046..6079
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(6148..6165)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6319..6907)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6399..6418)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7078..7869,7870..7938))
                     /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     7701..7720
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7939..8043)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8111..8129
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(8167..8189)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     8289..8308
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      join(8325..8787,1..702)
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 agtaagaact attcatagag tgaatcgaaa acaatacgaa aatgtaaaca tttcctatac
       61 gtagtatata gagacaaaat agaagaaacc gttcataatt ttctgaccaa tgaagaatca
      121 tcaacgctat cactttctgt tcacaaagta tgcgcaatcc acatcggtat agaatataat
      181 cggggatgcc tttatcttga aaaaatgcac ccgcagcttc gctagtaatc agtaaacgcg
      241 ggaagtggag tcaggctttt tttatggaag agaaaataga caccaaagta gccttcttct
      301 aaccttaacg gacctacagt gcaaaaagtt atcaagagac tgcattatag agcgcacaaa
      361 ggagaaaaaa agtaatctaa gatgctttgt tagaaaaata gcgctctcgg gatgcatttt
      421 tgtagaacaa aaaagaagta tagattcttt gttggtaaaa tagcgctctc gcgttgcatt
      481 tctgttctgt aaaaatgcag ctcagattct ttgtttgaaa aattagcgct ctcgcgttgc
      541 atttttgttt tacaaaaatg aagcacagat tcttcgttgg taaaatagcg ctttcgcgtt
      601 gcatttctgt tctgtaaaaa tgcagctcag attctttgtt tgaaaaatta gcgctctcgc
      661 gttgcatttt tgttctacaa aatgaagcac agatgcttcg ttgcttgcat gcctgcaggt
      721 cgagatccgg gatcgaagaa atgatggtaa atgaaatagg aaatcaagga gcatgaaggc
      781 aaaagacaaa tataagggtc gaacgaaaaa taaagtgaaa agtgttgata tgatgtattt
      841 ggctttgcgg cgccgaaaaa acgagtttac gcaattgcac aatcatgctg actctgtggc
      901 ggacccgcgc tcttgccggc ccggcgataa cgctgggcgt gaggctgtgc ccggcggagt
      961 tttttgcgcc tgcattttcc aaggtttacc ctgcgctaag gggcgagatt ggagaagcaa
     1021 taagaatgcc ggttggggtt gcgatgatga cgaccacgac aactggtgtc attatttaag
     1081 ttgccgaaag aacctgagtg catttgcaac atgagtatac tagaagaatg agccaagact
     1141 tgcgagacgc gagtttgccg gtggtgcgaa caatagagcg accatgacct tgaaggtgag
     1201 acgcgcataa ccgctagagt actttgaaga ggaaacagca atagggttgc taccagtata
     1261 aatagacagg tacatacaac actggaaatg gttgtctgtt tgagtacgct ttcaattcat
     1321 ttgggtgtgc actttattat gttacaatat ggaagggaac tttacacttc tcctatgcac
     1381 atatattaat taaagtccaa tgctagtaga gaaggggggt aacacccctc cgcgctcttt
     1441 tccgattttt ttctaaaccg tggaatattt cggatatcct tttgttgttt ccgggtgtac
     1501 aatatggact tcctcttttc tggcaaccaa acccatacat cgggattcct ataatacctt
     1561 cgttggtctc cctaacatgt aggtggcgga ggggagatat acaatagaac agataccaga
     1621 caagacataa tgggctaaac aagactacac caattacact gcctcattga tggtggtaca
     1681 taacgaacta atactgtagc cctagacttg atagccatca tcatatcgaa gtttcactac
     1741 cctttttcca tttgccatct attgaagtaa taataggcgc atgcaacttc ttttcttttt
     1801 ttttcttttc tctctccccc gttgttgtct caccatatcc gcaatgacaa aaaaatgatg
     1861 gaagacacta aaggaaaaaa ttaacgacaa agacagcacc aacagatgtc gttgttccag
     1921 agctgatgag gggtatctcg aagcacacga aactttttcc ttccttcatt cacgcacact
     1981 actctctaat gagcaacggt atacggcctt ccttccagtt acttgaattt gaaataaaaa
     2041 aaagtttgct gtcttgctat caagtataaa tagacctgca attattaatc ttttgtttcc
     2101 tcgtcattgt tctcgttccc tttcttcctt gtttcttttt ctgcacaata tttcaagcta
     2161 taccaagcat acaatcaact ccaagctttg caaagatgga taaagcggaa ttaattcccg
     2221 agcctccaaa aaagaagaga aaggtcgaat tgggtaccgc cgcctcggag ctccacttag
     2281 acggcgagga cgtggcgatg gcgcatgccg acgcgctaga cgatttcgat ctggacatgt
     2341 tgggggacgg ggattccccg ggtccgggat ttacccccca cgactccgcc ccctacggcg
     2401 ctctggatat ggccgacttc gagtttgagc agatgtttac cgatgccctt ggaattgacg
     2461 agtacggtgg gtttaatacg actcactata gggcgagcgc cgaagctagc gccaccatga
     2521 atggagctat aggaggtgac cttttgctca attttcctga catgtcggtc ctagagcgcc
     2581 aaagggctca cctcaagtac ctcaatccca cctttgattc tcctctcgcc ggcttctttg
     2641 ccgattcttc aatgattacc ggcggcgaga tggacagcta tctttcgact gccggtttga
     2701 atcttccgat gatgtacggt gagacgacgg tggaaggtga ttcaagactc tcaatttcgc
     2761 cggaaacgac gcttgggact ggaaatttca agaaacggaa gtttgataca gagactaagg
     2821 attgtaatga gaagaagaag aagatgacga tgaacagaga tgacctagta gaagaaggag
     2881 aagaagagaa gtcgaaaata acagagcaaa acaatgggag cacaaaaagc atcaagaaga
     2941 tgaaacacaa agccaagaaa gaagagaaca atttctctaa tgattcatct aaagtgacga
     3001 aggaattgga gaaaacggat tatattcatg ttcgtgcacg acgaggccaa gccactgata
     3061 gtcacagcat agcagaacga gttagaagag aaaagatcag tgagagaatg aagtttctac
     3121 aagatttggt tcctggatgc gacaagatca caggcaaagc agggatgctt gatgaaatca
     3181 ttaactatgt tcagtctctt cagagacaaa tcgagttctt atcgatgaaa ctagcaattg
     3241 tgaatccaag gccggatttt gatatggatg acatttttgc caaagaggtt gcctcaactc
     3301 caatgactgt ggtgccatct cctgaaatgg ttctttccgg ttattctcat gagatggttc
     3361 actctggtta ttctagtgag atggttaact ccggttacct tcatgtcaat ccaatgcagc
     3421 aagtgaatac cagttctgat ccattgtcat gcttcaacaa tggcgaagct ccttcgatgt
     3481 gggactctca tgtgcagaat ctctatggca atttaggagt accggtcatc gagctcgagc
     3541 tgcagatgaa tcgtagatac tgaaaaaccc cgcaagttca cttcaactgt gcatcgtgca
     3601 ccatctcaat ttctttcatt tatacatcgt tttgccttct tttatgtaac tatactcctc
     3661 taagtttcaa tcttggccat gtaacctctg atctatagaa ttttttaaat gactagaatt
     3721 aatgcccatc ttttttttgg acctaaattc ttcatgaaaa tatattacga gggcttattc
     3781 agaagctttg gacttcttcg ccagaggttt ggtcaagtct ccaatcaagg ttgtcggctt
     3841 gtctaccttg ccagaaattt acgaaaagat ggaaaagggt caaatcgttg gtagatacgt
     3901 tgttgacact tctaaataag cgaatttctt atgatttatg atttttatta ttaaataagt
     3961 tataaaaaaa ataagtgtat acaaatttta aagtgactct taggttttaa aacgaaaatt
     4021 cttattcttg agtaactctt tcctgtaggt caggttgctt tctcaggtat agcatgaggt
     4081 cgctcttatt gaccacacct ctaccggccg gtcgaaattc ccctacccta tgaacatatt
     4141 ccattttgta atttcgtgtc gtttctatta tgaatttcat ttataaagtt tatgtacaaa
     4201 tatcataaaa aaagagaatc tttttaagca aggattttct taacttcttc ggcgacagca
     4261 tcaccgactt cggtggtact gttggaacca cctaaatcac cagttctgat acctgcatcc
     4321 aaaacctttt taactgcatc ttcaatggcc ttaccttctt caggcaagtt caatgacaat
     4381 ttcaacatca ttgcagcaga caagatagtg gcgatagggt tgaccttatt ctttggcaaa
     4441 tctggagcag aaccgtggca tggttcgtac aaaccaaatg cggtgttctt gtctggcaaa
     4501 gaggccaagg acgcagatgg caacaaaccc aaggaacctg ggataacgga ggcttcatcg
     4561 gagatgatat caccaaacat gttgctggtg attataatac catttaggtg ggttgggttc
     4621 ttaactagga tcatggcggc agaatcaatc aattgatgtt gaaccttcaa tgtaggaaat
     4681 tcgttcttga tggtttcctc cacagttttt ctccataatc ttgaagaggc caaaacatta
     4741 gctttatcca aggaccaaat aggcaatggt ggctcatgtt gtagggccat gaaagcggcc
     4801 attcttgtga ttctttgcac ttctggaacg gtgtattgtt cactatccca agcgacacca
     4861 tcaccatcgt cttcctttct cttaccaaag taaatacctc ccactaattc tctgacaaca
     4921 acgaagtcag tacctttagc aaattgtggc ttgattggag ataagtctaa aagagagtcg
     4981 gatgcaaagt tacatggtct taagttggcg tacaattgaa gttctttacg gatttttagt
     5041 aaaccttgtt caggtctaac actacctgta ccccatttag gaccacccac agcacctaac
     5101 aaaacggcat cagccttctt ggaggcttcc agcgcctcat ctggaagtgg gacacctgta
     5161 gcttcgatag cagcaccacc aattaaatga ttttcgaaat cgaacttgac attggaacga
     5221 acatcagaaa tagctttaag aaccttaatg gcttcggctg tgatttcttg accaacgtgg
     5281 tcacctggca aaacgacgat cttcttaggg gcagacatta gaatggtata tccttgaaat
     5341 atatatatat attgctgaaa tgtaaaaggt aagaaaagtt agaaagtaag acgattgcta
     5401 accacctatt ggaaaaaaca ataggtcctt aaataatatt gtcaacttca agtattgtga
     5461 tgcaagcatt tagtcatgaa cgcttctcta ttctatatga aaagccggtt ccggcgctct
     5521 cacctttcct ttttctccca atttttcagt tgaaaaaggt atatgcgtca ggcgacctct
     5581 gaaattaaca aaaaatttcc agtcatcgaa tttgattctg tgcgatagcg cccctgtgtg
     5641 ttctcgttat gttgaggaaa aaaataatgg ttgctaagag attcgaactc ttgcatctta
     5701 cgatacctga gtattcccac agttggggga tctcgactct agctagagga tcaattcgta
     5761 atcatgtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     5821 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     5881 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgataact
     5941 tcgtataatg tatgctatac gaagttatta ggtctgaaga ggagtttacg tccagccaag
     6001 ctagcttggc tgcaggtcga gcggccgcga tccggaaccc ttaatataac ttcgtataat
     6061 gtatgctata cgaagttatc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6121 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     6181 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6241 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6301 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6361 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6421 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6481 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     6541 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6601 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     6661 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     6721 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     6781 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     6841 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     6901 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     6961 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     7021 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     7081 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     7141 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     7201 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     7261 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     7321 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     7381 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     7441 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     7501 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     7561 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     7621 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     7681 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     7741 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     7801 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     7861 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     7921 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     7981 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     8041 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     8101 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     8161 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     8221 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     8281 taactatgcg gcatcagagc agattgtact gagagtgcac cataacgcat ttaagcataa
     8341 acacgcacta tgccgttctt ctcatgtata tatatataca ggcaacacgc agatataggt
     8401 gcgacgtgaa cagtgagctg tatgtgcgca gctcgcgttg cattttcgga agcgctcgtt
     8461 ttcggaaacg ctttgaagtt cctattccga agttcctatt ctctagctag aaagtatagg
     8521 aacttcagag cgcttttgaa aaccaaaagc gctctgaaga cgcactttca aaaaaccaaa
     8581 aacgcaccgg actgtaacga gctactaaaa tattgcgaat accgcttcca caaacattgc
     8641 tcaaaagtat ctctttgcta tatatctctg tgctatatcc ctatataacc tacccatcca
     8701 cctttcgctc cttgaacttg catctaaact cgacctctac attttttatg tttatctcta
     8761 gtattactct ttagacaaaa aaattgt
//
