LOCUS       sequence-247336-        7396 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  LEU2 integrtion vector for VP16-CIB1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7396)
  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 7396)
  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: pMM0554
FEATURES             Location/Qualifiers
     source          1..7396
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        736..1402
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     1272..1295
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             1439..1459
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     1687..1706
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1687..1705
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1730..2239
                     /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      2789..2976
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2844..2868)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        3431..3838
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             3839..4933
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     CDS             complement(5661..6476)
                     /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(5661..6476)
                     /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     6384..6403
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6468..6487
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6701..7289
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7190..7209
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 caatacccag aagaacagta aaataaagca aggtacgtga aattaatatt tttaaatggt
       61 tctaaccgat gccgaagaac tgcgcagtcc ggttataacg tctgacatgt ccttttttga
      121 tttggaatcc aaccactcaa gtgactctgt tcatttactt tgcgaaaaat atacccacaa
      181 attgcccatc gaaagtgaat cgcaaaccac cttcagactg gcaccgacaa agcaaagatt
      241 atacagacag agtactttat acgtaccgtt aagtctcaag caaagggttt tcttatttac
      301 tgaacgggta aagagtatct gggccggctt gccaagatgc aaaccgaata agtatttcaa
      361 agttgcattt gccttagccg tcctgacacc attggctatt tggatatttt atattgactt
      421 tcgtgtacat tgatcacatc gactgttcta ttggcaaatg aaccacgggc attgactatt
      481 tttcaggtta ctactatata ttatcatcac gggcaaggat tgtaccctga attcgcatct
      541 agatggtaga gccacaaaca gccggtacaa gcaacgatct ccaggaccat ctgaatcatg
      601 cgcggatgac acgaactcac gacggcgatc acagacatta acccacagta cagacactgc
      661 gacaacgtgg caattcgtcg caataccgtc tcactgaact ggccgataat tgcagacgaa
      721 cgcagttcga gtttatcatt atcaatactg ccatttcaaa gaatacgtaa ataattaata
      781 gtagtgattt tcctaacttt atttagtcaa aaaattagcc ttttaattct gctgtaaccc
      841 gtacatgccc aaaatagggg gcgggttaca cagaatatat aacatcgtag gtgtctgggt
      901 gaacagttta ttcctggcat ccactaaata taatggagcc cgctttttaa gctggcatcc
      961 agaaaaaaaa agaatcccag caccaaaata ttgttttctt caccaaccat cagttcatag
     1021 gtccattctc ttagcgcaac tacagagaac aggggcacaa acaggcaaaa aacgggcaca
     1081 acctcaatgg agtgatgcaa cctgcctgga gtaaatgatg acacaaggca attgacccac
     1141 gcatgtatct atctcatttt cttacacctt ctattacctt ctgctctctc tgatttggaa
     1201 aaagctgaaa aaaaaggttg aaaccagttc cctgaaatta ttcccctact tgactaataa
     1261 gtatataaag acggtaggta ttgattgtaa ttctgtaaat ctatttctta aacttcttaa
     1321 attctacttt tatagttagt ctttttttta gttttaaaac accaagaact tagtttcgaa
     1381 taaacacaca taaacaaaca aaagatctat ggataaagcg gaattaattc ccgagcctcc
     1441 aaaaaagaag agaaaggtcg aattgggtac cgccgcctcg gagctccact tagacggcga
     1501 ggacgtggcg atggcgcatg ccgacgcgct agacgatttc gatctggaca tgttggggga
     1561 cggggattcc ccgggtccgg gatttacccc ccacgactcc gccccctacg gcgctctgga
     1621 tatggccgac ttcgagtttg agcagatgtt taccgatgcc cttggaattg acgagtacgg
     1681 tgggtttaat acgactcact atagggcgag cgccgaagct agcgccacca tgaatggagc
     1741 tataggaggt gaccttttgc tcaattttcc tgacatgtcg gtcctagagc gccaaagggc
     1801 tcacctcaag tacctcaatc ccacctttga ttctcctctc gccggcttct ttgccgattc
     1861 ttcaatgatt accggcggcg agatggacag ctatctttcg actgccggtt tgaatcttcc
     1921 gatgatgtac ggtgaaacga cggtggaagg tgattcaaga ctctcaattt cgccggaaac
     1981 gacgcttggg actggaaatt tcaagaaacg gaagtttgat acagagacta aggattgtaa
     2041 tgagaagaag aagaagatga cgatgaacag agatgaccta gtagaagaag gagaagaaga
     2101 gaagtcgaaa ataacagagc aaaacaatgg gagcacaaaa agcatcaaga agatgaaaca
     2161 caaagccaag aaagaagaga acaatttctc taatgattca tctaaagtga cgaaggaatt
     2221 ggagaaaacg gattatattc atgttcgtgc acgacgaggc caagccactg atagtcacag
     2281 catagcagaa cgagttagaa gagaaaagat cagtgagaga atgaagtttc tacaagattt
     2341 ggttcctgga tgcgacaaga tcacaggcaa agcagggatg cttgatgaaa tcattaacta
     2401 tgttcagtct cttcagagac aaatcgagtt cttatcgatg aaactagcaa ttgtgaatcc
     2461 aaggccggat tttgatatgg atgacatttt tgccaaagag gttgcctcaa ctccaatgac
     2521 tgtggtgcca tctcctgaaa tggttctttc cggttattct catgagatgg ttcactctgg
     2581 ttattctagt gagatggtta actccggtta ccttcatgtc aatccaatgc agcaagtgaa
     2641 taccagttct gatccattgt catgcttcaa caatggcgaa gctccttcga tgtgggactc
     2701 tcatgtgcag aatctctatg gcaatttagg agtaccggtc atcgagctcg agctgcagat
     2761 gaatcgtaga tacggatcct aactcgaggc gaatttctta tgatttatga tttttattat
     2821 taaataagtt ataaaaaaaa taagtgtata caaattttaa agtgactctt aggttttaaa
     2881 acgaaaattc ttattcttga gtaactcttt cctgtaggtc aggttgcttt ctcaggtata
     2941 gcatgaggtc gctcttattg accacacctc taccggcatg ccgagcaaat gcctgcaaat
     3001 cgctccccat ttcgctgcca atgagacgac ggggtcatca cggctcatca tgcgccaaac
     3061 aaatgtgtgc aatacacgct cggatgactg catgatgacc gcactgactg gggacagcag
     3121 atccacctaa gcctgtgaga gaagcagaca cccgacagat caaggcagtt aactagtgca
     3181 ctgcagtaca tcgaggagaa cttctagtat atctacatac ctaatattat tgccttatta
     3241 aaaatggaat cccaacaatt acatcaaaat ccacattctc ttcaaaatca attgtcctgt
     3301 acttccttgt tcatgtgtgt tcaaaaacgt tatatttata ggataattat actctatttc
     3361 tcaacaagta attggttgtt tggccgagcg gtctaaggcg cctgattcaa gaaatatctt
     3421 gaccgcagtt aactgtggga atactcaggt atcgtaagat gcaagagttc gaatctctta
     3481 gcaaccatta tttttttcct caacataacg agaacacaca ggggcgctat cgcacagaat
     3541 caaattcgat gactggaaat tttttgttaa tttcagaggt cgcctgacgc atataccttt
     3601 ttcaactgaa aaattgggag aaaaaggaaa ggtgagagcg ccggaaccgg cttttcatat
     3661 agaatagaga agcgttcatg actaaatgct tgcatcacaa tacttgaagt tgacaatatt
     3721 atttaaggac ctattgtttt ttccaatagg tggttagcaa tcgtcttact ttctaacttt
     3781 tcttaccttt tacatttcag caatatatat atatatattt caaggatata ccattctaat
     3841 gtctgcccct aagaagatcg tcgttttgcc aggtgaccac gttggtcaag aaatcacagc
     3901 cgaagccatt aaggttctta aagctatttc tgatgttcgt tccaatgtca agttcgattt
     3961 cgaaaatcat ttaattggtg gtgctgctat cgatgctaca ggtgttccac ttccagatga
     4021 ggcgctggaa gcctccaaga aggctgatgc cgttttgtta ggtgctgtgg gtggtcctaa
     4081 atggggaacc ggtagtgtta gacctgaaca aggtttacta aaaatccgta aagaacttca
     4141 attgtacgcc aacttaagac catgtaactt tgcatccgac tctcttttag acttatctcc
     4201 aatcaagcca caatttgcta aaggtactga cttcgttgtt gtcagagaat tagtgggagg
     4261 tatttacttt ggtaagagaa aggaggacga tggtgatggt gtcgcttggg atagtgaaca
     4321 atacaccgtt ccagaagtgc aaagaatcac aagaatggcc gctttcatgg ccctacaaca
     4381 tgagccacca ttgcctattt ggtccttgga taaagctaat gttttggcct cttcaagatt
     4441 atggagaaaa actgtggagg aaaccatcaa gaacgaattt cctacattga aggttcaaca
     4501 tcaattgatt gattctgccg ccatgatcct agttaagaac ccaacccacc taaatggtat
     4561 tataatcacc agcaacatgt ttggtgatat catctccgat gaagcctccg ttatcccagg
     4621 ttccttgggt ttgttgccat ctgcgtcctt ggcctctttg ccagacaaga acaccgcatt
     4681 tggtttgtac gaaccatgcc acggttctgc tccagatttg ccaaagaata aggtcaaccc
     4741 tatcgccact atcttgtctg ctgcaatgat gttgaaattg tcattgaact tgcctgaaga
     4801 aggtaaggcc attgaagatg cagttaaaaa ggttttggat gcaggtatca gaactggtga
     4861 tttaggtggt tccaacagta ccaccgaagt cggtgatgct gtcgccgaag aagttaagaa
     4921 aatccttgct taacctggag gacccttctc tttagactat tctactctta tgcacgtaaa
     4981 aaattctagg aaatatgtat taactaggag taaaataacc ggctagtggc attcatatag
     5041 ccgtctgttt acatctacat cacacatttc gagtgtatat ctcgcaacgt tggcgttaaa
     5101 taggcaggag tactcgtatc gcatgtcggt gcgacacgaa attacaaaat ggaatatgtt
     5161 catagggtag acgaaactat atacgcaatc tacatacatt tatcaagaag gagaaaaagg
     5221 aggatgtaaa ggaatacagg taagcaaatt gatactaatg gctcaacgtg ataaggaaaa
     5281 agaattgcac tttaacatta atattgacaa ggaggagggc accacacaaa aagttaggtg
     5341 taacagaaaa tcatgaaact atgattccta atttatatat tggaggattt tctctaaaaa
     5401 aaaaaaaata caacaaataa aaaacactca atgacctgac catttgatgg agtttaagtc
     5461 aataccttct tgaaccattt cccataatgg tgaaagttcc ctcaagaatt ttactctgtc
     5521 agaaacggcc ttaacgacgt agtcgacctc ctcttcagta ctaaatctac caataccaaa
     5581 tctgatggaa gaatgggcta atgcatcatc cttacccagc gccgagcggc cgcgtgttac
     5641 aaccaattaa ccaattctga ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta
     5701 ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa
     5761 aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc gattccgact
     5821 cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt atcaagtgag
     5881 aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagcttatg catttctttc
     5941 cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa
     6001 ccgttattca ttcgtgattg cgcctgagcg aggcgaaata cgcgatcgct gttaaaagga
     6061 caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata
     6121 ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc ggggatcgca
     6181 gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt cggaagaggc
     6241 ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt ggcaacgcta
     6301 cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa tcgatagatt
     6361 gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa atcagcatcc
     6421 atgttggaat ttaatcgcgg cctggagcaa gacgtttccc gttgaatatg gctcataaca
     6481 ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgatga tatattttta
     6541 tcttgtgcaa tgtaacatca gagattttga gacacaacgt ggctttgttg aataaatcga
     6601 acttttgctg agttgaagga tcagtcatga ccaaaatccc ttaacgtgag ttttcgttcc
     6661 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     6721 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     6781 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     6841 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     6901 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     6961 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     7021 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     7081 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     7141 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     7201 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     7261 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     7321 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     7381 ataaccgtgc ggccgc
//
