LOCUS       sequence-259402-        8792 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Overexpression plasmid for VP16-CIB1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8792)
  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 8792)
  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: pMM0520
FEATURES             Location/Qualifiers
     source          1..8792
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             46..66
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     294..313
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        294..312
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             337..846
                     /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      1740..1927
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1795..1819)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2044..3138)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIEATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRKLVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(3139..3544)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(3586..3602)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3586..3602)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3599..3621)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3610..3626
                     /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(3634..3640,3641..3658,3659..3664))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3679..3700
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    3755..3788
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    3866..3899
                     /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(3968..3985)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4139..4727)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4219..4238)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4898..5689,5690..5758))
                     /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     5521..5540
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5759..5863)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5931..5949
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5987..6009)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6109..6128
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      6145..7313
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        8087..8792
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
ORIGIN
        1 ccaagctttg caaagatgga taaagcggaa ttaattcccg agcctccaaa aaagaagaga
       61 aaggtcgaat tgggtaccgc cgcctcggag ctccacttag acggcgagga cgtggcgatg
      121 gcgcatgccg acgcgctaga cgatttcgat ctggacatgt tgggggacgg ggattccccg
      181 ggtccgggat ttacccccca cgactccgcc ccctacggcg ctctggatat ggccgacttc
      241 gagtttgagc agatgtttac cgatgccctt ggaattgacg agtacggtgg gtttaatacg
      301 actcactata gggcgagcgc cgaagctagc gccaccatga atggagctat aggaggtgac
      361 cttttgctca attttcctga catgtcggtc ctagagcgcc aaagggctca cctcaagtac
      421 ctcaatccca cctttgattc tcctctcgcc ggcttctttg ccgattcttc aatgattacc
      481 ggcggcgaga tggacagcta tctttcgact gccggtttga atcttccgat gatgtacggt
      541 gaaacgacgg tggaaggtga ttcaagactc tcaatttcgc cggaaacgac gcttgggact
      601 ggaaatttca agaaacggaa gtttgataca gagactaagg attgtaatga gaagaagaag
      661 aagatgacga tgaacagaga tgacctagta gaagaaggag aagaagagaa gtcgaaaata
      721 acagagcaaa acaatgggag cacaaaaagc atcaagaaga tgaaacacaa agccaagaaa
      781 gaagagaaca atttctctaa tgattcatct aaagtgacga aggaattgga gaaaacggat
      841 tatattcatg ttcgtgcacg acgaggccaa gccactgata gtcacagcat agcagaacga
      901 gttagaagag aaaagatcag tgagagaatg aagtttctac aagatttggt tcctggatgc
      961 gacaagatca caggcaaagc agggatgctt gatgaaatca ttaactatgt tcagtctctt
     1021 cagagacaaa tcgagttctt atcgatgaaa ctagcaattg tgaatccaag gccggatttt
     1081 gatatggatg acatttttgc caaagaggtt gcctcaactc caatgactgt ggtgccatct
     1141 cctgaaatgg ttctttccgg ttattctcat gagatggttc actctggtta ttctagtgag
     1201 atggttaact ccggttacct tcatgtcaat ccaatgcagc aagtgaatac cagttctgat
     1261 ccattgtcat gcttcaacaa tggcgaagct ccttcgatgt gggactctca tgtgcagaat
     1321 ctctatggca atttaggagt accggtcatc gagctcgagc tgcagatgaa tcgtagatac
     1381 tgaaaaaccc cgcaagttca cttcaactgt gcatcgtgca ccatctcaat ttctttcatt
     1441 tatacatcgt tttgccttct tttatgtaac tatactcctc taagtttcaa tcttggccat
     1501 gtaacctctg atctatagaa ttttttaaat gactagaatt aatgcccatc ttttttttgg
     1561 acctaaattc ttcatgaaaa tatattacga gggcttattc agaagctttg gacttcttcg
     1621 ccagaggttt ggtcaagtct ccaatcaagg ttgtcggctt gtctaccttg ccagaaattt
     1681 acgaaaagat ggaaaagggt caaatcgttg gtagatacgt tgttgacact tctaaataag
     1741 cgaatttctt atgatttatg atttttatta ttaaataagt tataaaaaaa ataagtgtat
     1801 acaaatttta aagtgactct taggttttaa aacgaaaatt cttattcttg agtaactctt
     1861 tcctgtaggt caggttgctt tctcaggtat agcatgaggt cgctcttatt gaccacacct
     1921 ctaccggccg gtcgaaattc ccctacccta tgaacatatt ccattttgta atttcgtgtc
     1981 gtttctatta tgaatttcat ttataaagtt tatgtacaaa tatcataaaa aaagagaatc
     2041 tttttaagca aggattttct taacttcttc ggcgacagca tcaccgactt cggtggtact
     2101 gttggaacca cctaaatcac cagttctgat acctgcatcc aaaacctttt taactgcatc
     2161 ttcaatggcc ttaccttctt caggcaagtt caatgacaat ttcaacatca ttgcagcaga
     2221 caagatagtg gcgatagggt tgaccttatt ctttggcaaa tctggagcag aaccgtggca
     2281 tggttcgtac aaaccaaatg cggtgttctt gtctggcaaa gaggccaagg acgcagatgg
     2341 caacaaaccc aaggaacctg ggataacgga ggcttcatcg gagatgatat caccaaacat
     2401 gttgctggtg attataatac catttaggtg ggttgggttc ttaactagga tcatggcggc
     2461 agaatcaatc aattgatgtt gaaccttcaa tgtaggaaat tcgttcttga tggtttcctc
     2521 cacagttttt ctccataatc ttgaagaggc caaaacatta gctttatcca aggaccaaat
     2581 aggcaatggt ggctcatgtt gtagggccat gaaagcggcc attcttgtga ttctttgcac
     2641 ttctggaacg gtgtattgtt cactatccca agcgacacca tcaccatcgt cttcctttct
     2701 cttaccaaag taaatacctc ccactaattt tctgacaaca acgaagtcag tacctttagc
     2761 aaattgtggc ttgattggag ataagtctaa aagagagtcg gatgcaaagt tacatggtct
     2821 taagttggcg tacaattgaa gttctttacg gatttttagt aaaccttgtt caggtctaac
     2881 actacctgta ccccatttag gaccacccac agcacctaac aaaacggcat cagccttctt
     2941 ggaggcttcc agcgcctcat ctggaagtgg gacacctgta gcttcgatag cagcaccacc
     3001 aattaaatga ttttcgaaat cgaacttgac attggaacga acatcagaaa tagctttaag
     3061 aaccttaatg gcttcggctg tgatttcttg accaacgtgg tcacctggca aaacgacgat
     3121 cttcttaggg gcagacatta gaatggtata tccttgaaat atatatatat attgctgaaa
     3181 tgtaaaaggt aagaaaagtt agaaagtaag acgattgcta accacctatt ggaaaaaaca
     3241 ataggtcctt aaataatatt gtcaacttca agtattgtga tgcaagcatt tagtcatgaa
     3301 cgcttctcta ttctatatga aaagccggtt ccggcgctct cacctttcct ttttctccca
     3361 atttttcagt tgaaaaaggt atatgcgtca ggcgacctct gaaattaaca aaaaatttcc
     3421 agtcatcgaa tttgattctg tgcgatagcg cccctgtgtg ttctcgttat gttgaggaaa
     3481 aaaataatgg ttgctaagag attcgaactc ttgcatctta cgatacctga gtattcccac
     3541 agttggggga tctcgactct agctagagga tcaattcgta atcatgtcat agctgtttcc
     3601 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     3661 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     3721 cgctttccag tcgggaaacc tgtcgtgcca gctgataact tcgtataatg tatgctatac
     3781 gaagttatta ggtctgaaga ggagtttacg tccagccaag ctagcttggc tgcaggtcga
     3841 gcggccgcga tccggaaccc ttaatataac ttcgtataat gtatgctata cgaagttatc
     3901 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     3961 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     4021 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     4081 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     4141 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     4201 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     4261 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     4321 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     4381 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     4441 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4501 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4561 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4621 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4681 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4741 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     4801 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     4861 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     4921 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     4981 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     5041 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     5101 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     5161 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     5221 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     5281 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     5341 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     5401 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     5461 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     5521 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     5581 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     5641 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     5701 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     5761 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     5821 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     5881 tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta
     5941 tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc
     6001 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc
     6061 agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc
     6121 agattgtact gagagtgcac cataacgcat ttaagcataa acacgcacta tgccgttctt
     6181 ctcatgtata tatatatata tacaggcaac acgcagatat aggtgcgacg tgaacagtga
     6241 gctgtatgtg cgcagctcgc gttgcatttt cggaagcgct cgttttcgga aacgctttga
     6301 agttcctatt ccgaagttcc tattctctag ctagaaagta taggaacttc agagcgcttt
     6361 tgaaaaccaa aagcgctctg aagacgcact ttcaaaaaac caaaaacgca ccggactgta
     6421 acgagctact aaaatattgc gaataccgct tccacaaaca ttgctcaaaa gtatctcttt
     6481 gctatatatc tctgtgctat atccctatat aacctaccca tccacctttc gctccttgaa
     6541 cttgcatcta aactcgacct ctacattttt tatgtttatc tctagtatta ctctttagac
     6601 aaaaaaattg tagtaagaac tattcataga gtgaatcgaa aacaatacga aaatgtaaac
     6661 atttcctata cgtagtatat agagacaaaa tagaagaaac cgttcataat tttctgacca
     6721 atgaagaatc atcaacgcta tcactttctg ttcacaaagt atgcgcaatc cacatcggta
     6781 tagaatataa tcggggatgc ctttatcttg aaaaaatgca cccgcagctt cgctagtaat
     6841 cagtaaacgc gggaagtgga gtcaggcttt ttttatggaa gagaaaatag acaccaaagt
     6901 agccttcttc taaccttaac ggacctacag tgcaaaaagt tatcaagaga ctgcattata
     6961 gagcgcacaa aggagaaaaa aagtaatcta agatgctttg ttagaaaaat agcgctctcg
     7021 ggatgcattt ttgtagaaca aaaaagaagt atagattctt tgttggtaaa atagcgctct
     7081 cgcgttgcat ttctgttctg taaaaatgca gctcagattc tttgtttgaa aaattagcgc
     7141 tctcgcgttg catttttgtt ttacaaaaat gaagcacaga ttcttcgttg gtaaaatagc
     7201 gctttcgcgt tgcatttctg ttctgtaaaa atgcagctca gattctttgt ttgaaaaatt
     7261 agcgctctcg cgttgcattt ttgttctaca aaatgaagca cagatgcttc gttgcttgca
     7321 tgcctgcagg tcgagatccg ggatcgaaga aatgatggta aatgaaatag gaaatcaagg
     7381 agcatgaagg caaaagacaa atataagggt cgaacgaaaa ataaagtgaa aagtgttgat
     7441 atgatgtatt tggctttgcg gcgccgaaaa aacgagttta cgcaattgca caatcatgct
     7501 gactctgtgg cggacccgcg ctcttgccgg cccggcgata acgctgggcg tgaggctgtg
     7561 cccggcggag ttttttgcgc ctgcattttc caaggtttac cctgcgctaa ggggcgagat
     7621 tggagaagca ataagaatgc cggttggggt tgcgatgatg acgaccacga caactggtgt
     7681 cattatttaa gttgccgaaa gaacctgagt gcatttgcaa catgagtata ctagaagaat
     7741 gagccaagac ttgcgagacg cgagtttgcc ggtggtgcga acaatagagc gaccatgacc
     7801 ttgaaggtga gacgcgcata accgctagag tactttgaag aggaaacagc aatagggttg
     7861 ctaccagtat aaatagacag gtacatacaa cactggaaat ggttgtctgt ttgagtacgc
     7921 tttcaattca tttgggtgtg cactttatta tgttacaata tggaagggaa ctttacactt
     7981 ctcctatgca catatattaa ttaaagtcca atgctagtag agaagggggg taacacccct
     8041 ccgcgctctt ttccgatttt tttctaaacc gtggaatatt tcggatatcc ttttgttgtt
     8101 tccgggtgta caatatggac ttcctctttt ctggcaacca aacccataca tcgggattcc
     8161 tataatacct tcgttggtct ccctaacatg taggtggcgg aggggagata tacaatagaa
     8221 cagataccag acaagacata atgggctaaa caagactaca ccaattacac tgcctcattg
     8281 atggtggtac ataacgaact aatactgtag ccctagactt gatagccatc atcatatcga
     8341 agtttcacta ccctttttcc atttgccatc tattgaagta ataataggcg catgcaactt
     8401 cttttctttt ttttttcttt tctctctccc ccgttgttgt ctcaccatat ccgcaatgac
     8461 aaaaaaatga tggaagacac taaaggaaaa aattaacgac aaagacagca ccaacagatg
     8521 tcgttgttcc agagctgatg aggggtatct cgaagcacac gaaacttttt ccttccttca
     8581 ttcacgcaca ctactctcta atgagcaacg gtatacggcc ttccttccag ttacttgaat
     8641 ttgaaataaa aaaaagtttg ctgtcttgct atcaagtata aatagacctg caattattaa
     8701 tcttttgttt cctcgtcatt gttctcgttc cctttcttcc ttgtttcttt ttctgcacaa
     8761 tatttcaagc tataccaagc atacaatcaa ct
//
