LOCUS       sequence-259470-        9147 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Control for optogenetic machinery 
            (pMEDIUM-VP16-CIB1/pMEDIUM-ZCRY2PHR) integration effects.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9147)
  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 9147)
  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: pMM0652
FEATURES             Location/Qualifiers
     source          1..9147
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             222..1025
                     /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"
     CDS             complement(1667..2482)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1667..2482)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2390..2409
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2474..2493
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2707..3295
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3196..3215
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             4711..4731
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     4959..4978
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4959..4977
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5002..5511
                     /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"
     CDS             7031..7051
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7334..8827
                     /codon_start=1
                     /product="N-terminal photolyase homology region of 
                     Arabidopsis thaliana cryptochrome 2"
                     /label=CRY2PHR
                     /note="binds to CIB1 upon exposure to blue light (Kennedy 
                     et al., 2010)"
                     /translation="MKMDKKTIVWFRRDLRIEDNPALAAAAHEGSVFPVFIWCPEEEGQ
                     FYPGRASRWWMKQSLAHLSQSLKALGSDLTLIKTHNTISAILDCIRVTGATKVVFNHLY
                     DPVSLVRDHTVKEKLVERGISVQSYNGDLLYEPWEIYCEKGKPFTSFNSYWKKCLDMSI
                     ESVMLPPPWRLMPITAAAEAIWACSIEELGLENEAEKPSNALLTRAWSPGWSNADKLLN
                     EFIEKQLIDYAKNSKKVVGNSTSLLSPYLHFGEISVRHVFQCARMKQIIWARDKNSEGE
                     ESADLFLRGIGLREYSRYICFNFPFTHEQSLLSHLRFFPWDADVDKFKAWRQGRTGYPL
                     VDAGMRELWATGWMHNRIRVIVSSFAVKFLLLPWKWGMKYFWDTLLDADLECDILGWQY
                     ISGSIPDGHELDRLDNPALQGAKYDPEGEYIRQWLPELARLPTEWIHHPWDAPLTVLKA
                     SGVELGTNYAKPIVDIDTARELLAKAISRTREAQIMIGAA"
ORIGIN
        1 agcagttaca gagatgttac gaaccactag tgcactgcag tacacggttt ccttgaaatt
       61 tttttgattc ggtaatctcc gaacagaagg aagaacgaag gaaggagcac agacttagat
      121 tggtatatat acgcatatgt agtgttgaag aaacatgaaa ttgcccagta ttcttaaccc
      181 aactgcacag aacaaaaacg tgcaggaaac gaagataaat catgtcgaaa gctacatata
      241 aggaacgtgc tgctactcat cctagtcctg ttgctgccaa gctatttaat atcatgcacg
      301 aaaagcaaac aaacttgtgt gcttcattgg atgttcgtac caccaaggaa ttactggagt
      361 tagttgaagc attaggtccc aaaatttgtt tactaaaaac acatgtggat atcttgactg
      421 atttttccat ggagggcaca gttaagccgc taaaggcatt atccgccaag tacaattttt
      481 tactcttcga ggacagaaaa tttgctgaca ttggtaatac agtcaaattg cagtactctg
      541 cgggtgtata cagaatagca gaatgggcag acattacgaa tgcacacggt gtggtgggcc
      601 caggtattgt tagcggtttg aagcaggcgg cagaagaagt aacaaaggaa cctagaggcc
      661 ttttgatgtt agcagaattg tcatgcaagg gctccctatc tactggagaa tatactaagg
      721 gtactgttga cattgcgaag agcgacaaag attttgttat cggctttatt gctcaaagag
      781 acatgggtgg aagagatgaa ggttacgatt ggttgattat gacacccggt gtgggtttag
      841 atgacaaggg agatgcattg ggtcaacagt atagaaccgt ggatgatgtg gtttctacag
      901 gatctgacat tattattgtt ggaagaggac tatttgcaaa gggaagggat gctaaggtag
      961 agggtgaacg ttacagaaaa gcaggctggg aagcatattt gagaagatgc ggccagcaaa
     1021 actaaaaaac tgtattataa gtaaatgcat gtatactaaa ctcacaaatt agagcttcaa
     1081 tttaattata tcagttatta cccgagtaga gcacttgaat ccactgcccc gggaatctcg
     1141 gtcgtaatga tttctataat gacgaaaaaa aaaaaattgg aaagaaaaag cttcatggcc
     1201 tttataaaaa ggaactatcc aatacctcgc cagaaccaag taacagtatt ttacggggca
     1261 caaatcaaga acaataagac aggactgtaa agatggacgc attgaactcc aaagaacaac
     1321 aagagttcca aaaagtagtg gaacaaaagc aaatgaagga tttcatgcgt ttgtactcta
     1381 atctggtaga aagatgtttc acagactgtg tcaatgactt cacaacatca aagctaacca
     1441 ataaggaaca aacatgcatc atgaagtgct cagaaaagtt cttgaagcat agcgaacgtg
     1501 tagggcagcg tttccaagaa caaaacgctg ccttgggaca aggcttgggc cgataaggtg
     1561 tactggcgta tatatatcta attatgtatc tctggtgtag cccattttta gcatgtaaat
     1621 ataaagaccg agcggccgcg tgttacaacc aattaaccaa ttctgattag aaaaactcat
     1681 cgagcatcaa atgaaactgc aatttattca tatcaggatt atcaatacca tatttttgaa
     1741 aaagccgttt ctgtaatgaa ggagaaaact caccgaggca gttccatagg atggcaagat
     1801 cctggtatcg gtctgcgatt ccgactcgtc caacatcaat acaacctatt aatttcccct
     1861 cgtcaaaaat aaggttatca agtgagaaat caccatgagt gacgactgaa tccggtgaga
     1921 atggcaaaag cttatgcatt tctttccaga cttgttcaac aggccagcca ttacgctcgt
     1981 catcaaaatc actcgcatca accaaaccgt tattcattcg tgattgcgcc tgagcgaggc
     2041 gaaatacgcg atcgctgtta aaaggacaat tacaaacagg aatcgaatgc aaccggcgca
     2101 ggaacactgc cagcgcatca acaatatttt cacctgaatc aggatattct tctaatacct
     2161 ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca tgcatcatca ggagtacgga
     2221 taaaatgctt gatggtcgga agaggcataa attccgtcag ccagtttagt ctgaccatct
     2281 catctgtaac atcattggca acgctacctt tgccatgttt cagaaacaac tctggcgcat
     2341 cgggcttccc atacaatcga tagattgtcg cacctgattg cccgacatta tcgcgagccc
     2401 atttataccc atataaatca gcatccatgt tggaatttaa tcgcggcctg gagcaagacg
     2461 tttcccgttg aatatggctc ataacacccc ttgtattact gtttatgtaa gcagacagtt
     2521 ttattgttca tgatgatata tttttatctt gtgcaatgta acatcagaga ttttgagaca
     2581 caacgtggct ttgttgaata aatcgaactt ttgctgagtt gaaggatcag tcatgaccaa
     2641 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2701 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2761 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2821 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     2881 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     2941 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     3001 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3061 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3121 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3181 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3241 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3301 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     3361 tcctgcgtta tcccctgatt ctgtggataa ccgtgcggcc gccaatgcta aattcgagtg
     3421 aaacacagga agatcagaaa atcctcattt catccatatt aacaataatt tcaaatgttt
     3481 atttgcatta tttgaaacta ggcaagacaa gcaacgaaac gtttttgaaa attttgagta
     3541 ttttcaataa atttgtagag gactcagata ttgaaaaaaa gctacagcaa ttaatacttg
     3601 ataagaagag tattgagaag ggcaacggtt catcatctca tggatctgca catgaacaaa
     3661 caccagagtc aaacgacgtt gaaattgagg ctactgcgcc aattgatgac aatacagacg
     3721 atgataacaa accgaagtta tctgatgtag aaaaggatta aagatgctaa gagatagtga
     3781 tgatatttca taaataatgt aattctatat atgttaatta ccttttttgc gaggcatatt
     3841 tatggtgaag gataagtttt gaccatcaaa gaaggttaat gtggctgtgg tttcagggtc
     3901 cataaagccc acatggataa cattacccct gaattcgcat ctagactgaa ctggccgata
     3961 attgcagacg aacgaagagg atgtccaata ttttttttaa ggaataagga tacttcaaga
     4021 ctagattccc ccctgcattc ccatcagaac cgtaaacctt ggcgctttcc ttgggaagta
     4081 ttcaagaagt gccttgtccg gtttctgtgg ctcacaaacc agcgcgcccg atatggcttt
     4141 cttttcactt atgaatgtac cagtacggga caattagaac gctcctgtaa caatctcttt
     4201 gcaaatgtgg ggttacattc taaccatgtc acactgctga cgaaattcaa agtaaaaaaa
     4261 aatgggacca cgtcttgaga acgatagatt ttctttattt tacattgaac agtcgttgtc
     4321 tcagcgcgct ttatgttttc attcatactt catattataa aataacaaaa gaagaatttc
     4381 atattcacgc ccaagaaatc aggctgcttt ccaaatgcaa ttgacacttc attagccatc
     4441 acacaaaact ctttcttgct ggagcttctt ttaaaaaaga cctcagtaca ccaaacacgt
     4501 tacccgacct cgttatttta cgacaactat gataaaattc tgaagaaaaa ataaaaaaat
     4561 tttcatactt cttgctttta tttaaaccat tgaatgattt cttttgaaca aaactacctg
     4621 tttcaccaaa ggaaatagaa agaaaaaatc aattagaaga aaacaaaaaa caaaagatct
     4681 atggataaag cggaattaat tcccgagcct ccaaaaaaga agagaaaggt cgaattgggt
     4741 accgccgcct cggagctcca cttagacggc gaggacgtgg cgatggcgca tgccgacgcg
     4801 ctagacgatt tcgatctgga catgttgggg gacggggatt ccccgggtcc gggatttacc
     4861 ccccacgact ccgcccccta cggcgctctg gatatggccg acttcgagtt tgagcagatg
     4921 tttaccgatg cccttggaat tgacgagtac ggtgggttta atacgactca ctatagggcg
     4981 agcgccgaag ctagcgccac catgaatgga gctataggag gtgacctttt gctcaatttt
     5041 cctgacatgt cggtcctaga gcgccaaagg gctcacctca agtacctcaa tcccaccttt
     5101 gattctcctc tcgccggctt ctttgccgat tcttcaatga ttaccggcgg cgagatggac
     5161 agctatcttt cgactgccgg tttgaatctt ccgatgatgt acggtgaaac gacggtggaa
     5221 ggtgattcaa gactctcaat ttcgccggaa acgacgcttg ggactggaaa tttcaagaaa
     5281 cggaagtttg atacagagac taaggattgt aatgagaaga agaagaagat gacgatgaac
     5341 agagatgacc tagtagaaga aggagaagaa gagaagtcga aaataacaga gcaaaacaat
     5401 gggagcacaa aaagcatcaa gaagatgaaa cacaaagcca agaaagaaga gaacaatttc
     5461 tctaatgatt catctaaagt gacgaaggaa ttggagaaaa cggattatat tcatgttcgt
     5521 gcacgacgag gccaagccac tgatagtcac agcatagcag aacgagttag aagagaaaag
     5581 atcagtgaga gaatgaagtt tctacaagat ttggttcctg gatgcgacaa gatcacaggc
     5641 aaagcaggga tgcttgatga aatcattaac tatgttcagt ctcttcagag acaaatcgag
     5701 ttcttatcga tgaaactagc aattgtgaat ccaaggccgg attttgatat ggatgacatt
     5761 tttgccaaag aggttgcctc aactccaatg actgtggtgc catctcctga aatggttctt
     5821 tccggttatt ctcatgagat ggttcactct ggttattcta gtgagatggt taactccggt
     5881 taccttcatg tcaatccaat gcagcaagtg aataccagtt ctgatccatt gtcatgcttc
     5941 aacaatggcg aagctccttc gatgtgggac tctcatgtgc agaatctcta tggcaattta
     6001 ggagtaccgg tcatcgagct cgagctgcag atgaatcgta gatacggatc ctaactcgag
     6061 agcttttgat taagccttct agtccaaaaa acacgttttt ttgtcattta tttcattttc
     6121 ttagaatagt ttagtttatt cattttatag tcacgaatgt tttatgattc tatatagggt
     6181 tgcaaacaag catttttcat tttatgttaa aacaatttca ggtttacctt ttattctgct
     6241 tgtggtgacg cgtgtatccg cccgctcttt tggtcaccca tgtatgctgc caaaccagat
     6301 gtcaacacag ctacaacgaa gaggatgtcc aatatttttt ttaaggaata aggatacttc
     6361 aagactagat tcccccctgc attcccatca gaaccgtaaa ccttggcgct ttccttggga
     6421 agtattcaag aagtgccttg tccggtttct gtggctcaca aaccagcgcg cccgatatgg
     6481 ctttcttttc acttatgaat gtaccagtac gggacaatta gaacgctcct gtaacaatct
     6541 ctttgcaaat gtggggttac attctaacca tgtcacactg ctgacgaaat tcaaagtaaa
     6601 aaaaaatggg accacgtctt gagaacgata gattttcttt attttacatt gaacagtcgt
     6661 tgtctcagcg cgctttatgt tttcattcat acttcatatt ataaaataac aaaagaagaa
     6721 tttcatattc acgcccaaga aatcaggctg ctttccaaat gcaattgaca cttcattagc
     6781 catcacacaa aactctttct tgctggagct tcttttaaaa aagacctcag tacaccaaac
     6841 acgttacccg acctcgttat tttacgacaa ctatgataaa attctgaaga aaaaataaaa
     6901 aaattttcat acttcttgct tttatttaaa ccattgaatg atttcttttg aacaaaacta
     6961 cctgtttcac caaaggaaat agaaagaaaa aatcaattag aagaaaacaa aaaacaaaag
     7021 atctatggcc cccaagaaga agaggaaggt tggtattcac gggggtaccc gcccatatgc
     7081 ttgccctgtc gagtcctgcg atcgccgctt ttctcgctcg gatgagctta cccgccatat
     7141 ccgcatccat accggtcaga agcccttcca gtgtcgaatc tgcatgcgta acttcagtcg
     7201 tagtgaccac cttaccaccc acatccgcac ccacacaggc gagaagcctt ttgcctgtga
     7261 catttgtggg aggaagtttg ccaggagtga tgaacgcaag aggcatacca aaatccatac
     7321 aggtgctagc ttcatgaaga tggacaaaaa gactatagtt tggtttagaa gagatctaag
     7381 gattgaggat aatcctgcat tagcagcagc tgctcacgaa ggatctgttt ttcctgtctt
     7441 catttggtgt cctgaagaag aaggacagtt ttatcctgga agagcttcaa gatggtggat
     7501 gaaacaatca cttgctcact tatctcaatc cttgaaggct cttggatctg acctcacttt
     7561 aatcaaaacc cacaacacga tttcagcgat cttggattgt atccgcgtta ccggtgctac
     7621 aaaagtcgtc tttaaccacc tctatgatcc tgtttcgtta gttcgggacc ataccgtaaa
     7681 ggagaagctg gtggaacgtg ggatctctgt gcaaagctac aatggagatc tattgtatga
     7741 accgtgggag atatactgcg aaaagggcaa accttttacg agtttcaatt cttactggaa
     7801 gaaatgctta gatatgtcga ttgaatccgt tatgcttcct cctccttggc ggttgatgcc
     7861 aataactgca gcggctgaag cgatttgggc gtgttcgatt gaagaactag ggctggagaa
     7921 tgaggccgag aaaccgagca atgcgttgtt aactagagct tggagtccag gatggagcaa
     7981 tgctgataag ttactaaatg agttcatcga gaagcagttg atagattatg caaagaacag
     8041 caagaaagtt gttgggaatt ctacttcact actttctccg tatctccatt tcggggaaat
     8101 aagcgtcaga cacgttttcc agtgtgcccg gatgaaacaa attatatggg caagagataa
     8161 gaacagtgaa ggagaagaaa gtgcagatct ttttcttagg ggaatcggtt taagagagta
     8221 ttctcggtat atatgtttca acttcccgtt tactcacgag caatcgttgt tgagtcatct
     8281 tcggtttttc ccttgggatg ctgatgttga taagttcaag gcctggagac aaggcaggac
     8341 cggttatccg ttggtggatg ccggaatgag agagctttgg gctaccggat ggatgcataa
     8401 cagaataaga gtgattgttt caagctttgc tgtgaagttt cttctccttc catggaaatg
     8461 gggaatgaag tatttctggg atacactttt ggatgctgat ttggaatgtg acatccttgg
     8521 ctggcagtat atctctggga gtatccccga tggccacgag cttgatcgct tggacaatcc
     8581 cgcgttacaa ggcgccaaat atgacccaga aggtgagtac ataaggcaat ggcttcccga
     8641 gcttgcgaga ttgccaactg aatggatcca tcatccatgg gacgctcctt taaccgtact
     8701 caaagcttct ggtgtggaac tcggaacaaa ctatgcgaaa cccattgtag acatcgacac
     8761 agctcgtgag ctactagcta aagctatttc aagaacccgt gaagcacaga tcatgatcgg
     8821 agcagcagcc cggggatcct aactcgaggc caattggtgc ggcaattgat aataacgaaa
     8881 atgtctttta atgatctggg tataatgagg aattttccga acgtttttac tttatatata
     8941 tatatacatg taacatatat tctatacgct atagagaaag gaaatttttc aattaaaaaa
     9001 aaaatagaga aagagtttca cttcttgatt atcgctaaca ctaatggttg aagtactgct
     9061 actttaattt tatgctggat gcacacactg gcttaagatg acaacgacta gagatctatg
     9121 tgaggatcct aactcgagat cgagctg
//
