LOCUS       sequence-96591-e        5983 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Contains a promoter of two gal binding sites coupled with a core 
            promoter derived from native Leucine promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5983)
  AUTHORS   Blazeck J, Garg R, Reed B, Alper HS
  TITLE     Controlling promoter strength and regulation in Saccharomyces 
            cerevisiae using synthetic hybrid promoters.
  JOURNAL   Biotechnol Bioeng. 2012 Nov;109(11):2884-95. doi: 10.1002/bit.24552.
            Epub 2012 May 17.
  PUBMED    22565375
REFERENCE   2  (bases 1 to 5983)
  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"; doi: "10.1002/bit.24552";
            journalName: "Biotechnol Bioeng"; date: "2012-11"; volume: "109"; 
            issue: "11"; pages: "2884-95"
COMMENT     Sequence Label: Gal4pBS24-Pleum
FEATURES             Location/Qualifiers
     source          1..5983
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     136..153
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(306..311,312..329,330..336)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    344..360
                     /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     349..371
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     368..384
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     368..384
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     403..423
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        405..423
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             649..1365
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1368..1384)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1369..1385)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(1403..1421)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(1635..1654)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1636..1654)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1664..1681)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1664..1680)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1673..1695)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1825..2280
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1912..1931)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2122..2143
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2411..3214)
                     /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(3215..3435)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3461..3480)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3580..3602
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3641..3659)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    3701..4204
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        4241..4345
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4346..4414,4415..5206)
                     /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(4564..4583)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5377..5965
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5866..5885
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
       61 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
      121 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
      181 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
      241 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt acctcactca ttaggcaccc
      301 caggctttac actttatgct tccggctcct atgttgtgtg gaattgtgag cggataacaa
      361 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa
      421 agggaacaaa agctggagct ccgggatcct tggcgcgccc cttaattaac ggaagcttgg
      481 cgacagccac ggaagactct cctccggttt aaaccaatat tatttaagga cctattgttt
      541 tttccaatag gtggttagca atcgtcttac tttctaactt ttcttacctt ttacatttca
      601 gcaatatata tatatatatt tcaaggatat accattctat ctagaaaaat gtctaaaggt
      661 gaagaattat tcactggtgt tgtcccaatt ttggttgaat tagatggtga tgttaatggt
      721 cacaaatttt ctgtctccgg tgaaggtgaa ggtgatgcta cttacggtaa attgacctta
      781 aaatttattt gtactactgg taaattgcca gttccatggc caaccttagt cactacttta
      841 ggttatggtt tgatgtgttt tgctagatac ccagatcata tgaaacaaca tgactttttc
      901 aagtctgcca tgccagaagg ttatgttcaa gaaagaacta tttttttcaa agatgacggt
      961 aactacaaga ccagagctga agtcaagttt gaaggtgata ccttagttaa tagaatcgaa
     1021 ttaaaaggta ttgattttaa agaagatggt aacattttag gtcacaaatt ggaatacaac
     1081 tataactctc acaatgttta catcatggct gacaaacaaa agaatggtat caaagttaac
     1141 ttcaaaatta gacacaacat tgaagatggt tctgttcaat tagctgacca ttatcaacaa
     1201 aatactccaa ttggtgatgg tccagtcttg ttaccagaca accattactt atcctatcaa
     1261 tctgccttat ccaaagatcc aaacgaaaag agagaccaca tggtcttgtt agaatttgtt
     1321 actgctgctg gtattaccca tggtatggat gaattgtaca aataaatcga taccgtcgac
     1381 ctcgagtcat gtaattagtt atgtcacgct tacattcacg ccctcctccc acatccgctc
     1441 taaccgaaaa ggaaggagtt agacaacctg aagtctaggt ccctatttat tttttttaat
     1501 agttatgtta gtattaagaa cgttatttat atttcaaatt tttctttttt ttctgtacaa
     1561 acgcgtgtac gcatgtaaca ttatactgaa aaccttgctt gagaaggttt tgggacgctc
     1621 gggtacccaa ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac
     1681 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc
     1741 ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     1801 gcagcctgaa tggcgaatgg cgcgacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     1861 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     1921 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     1981 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     2041 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     2101 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     2161 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     2221 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     2281 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tagggtaata
     2341 actgatataa ttaaattgaa gctctaattt gtgagtttag tatacatgca tttacttata
     2401 atacagtttt ttagttttgc tggccgcatc ttctcaaata tgcttcccag cctgcttttc
     2461 tgtaacgttc accctctacc ttagcatccc ttccctttgc aaatagtcct cttccaacaa
     2521 taataatgtc agatcctgta gagaccacat catccacggt tctatactgt tgacccaatg
     2581 cgtctccctt gtcatctaaa cccacaccgg gtgtcataat caaccaatcg taaccttcat
     2641 ctcttccacc catgtctctt tgagcaataa agccgataac aaaatctttg tcgctcttcg
     2701 caatgtcaac agtaccctta gtatattctc cagtagatag ggagcccttg catgacaatt
     2761 ctgctaacat caaaaggcct ctaggttcct ttgttacttc ttctgccgcc tgcttcaaac
     2821 cgctaacaat acctgggccc accacaccgt gtgcattcgt aatgtctgcc cattctgcta
     2881 ttctgtatac acccgcagag tactgcaatt tgactgtatt accaatgtca gcaaattttc
     2941 tgtcttcgaa gagtaaaaaa ttgtacttgg cggataatgc ctttagcggc ttaactgtgc
     3001 cctccatgga aaaatcagtc aagatatcca catgtgtttt tagtaaacaa attttgggac
     3061 ctaatgcttc aactaactcc agtaattcct tggtggtacg aacatccaat gaagcacaca
     3121 agtttgtttg cttttcgtgc atgatattaa atagcttggc agcaacagga ctaggatgag
     3181 tagcagcacg ttccttatat gtagctttcg acatgattta tcttcgtttc ctgcaggttt
     3241 ttgttctgtg cagttgggtt aagaatactg ggcaatttca tgtttcttca acactacata
     3301 tgcgtatata taccaatcta agtctgtgct ccttccttcg ttcttccttc tgttcggaga
     3361 ttaccgaatc aaaaaaattt caaagaaacc gaaatcaaaa aaaagaataa aaaaaaaatg
     3421 atgaattgaa ttgaaaagct gtggtatggt gcactctcag tacaatctgc tctgatgccg
     3481 catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     3541 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     3601 ggttttcacc gtcatcaccg aaacgcgcga agacgaaagg gcctcgtgat acgcctattt
     3661 ttataggtta atgtcatgat aataatggtt tcttaggacg gatcgcttgc ctgtaactta
     3721 cacgcgcctc gtatctttta atgatggaat aatttgggaa tttactctgt gtttatttat
     3781 ttttatgttt tgtatttgga ttttagaaag taaataaaga aggtagaaga gttacggaat
     3841 gaagaaaaaa aaataaacaa aggtttaaaa aatttcaaca aaaagcgtac tttacatata
     3901 tatttattag acaagaaaag cagattaaat agatatacat tcgattaacg ataagtaaaa
     3961 tgtaaaatca caggattttc gtgtgtggtc ttctacacag acaagatgaa acaattcggc
     4021 attaatacct gagagcagga agagcaagat aaaaggtagt atttgttggc gatcccccta
     4081 gagtctttta catcttcgga aaacaaaaac tattttttct ttaatttctt tttttacttt
     4141 ctatttttaa tttatatatt tatattaaaa aatttaaatt ataattattt ttatagcacg
     4201 tgatgaaaag gacccaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     4261 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     4321 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     4381 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     4441 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     4501 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     4561 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     4621 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     4681 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     4741 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     4801 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     4861 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     4921 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     4981 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     5041 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     5101 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     5161 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     5221 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     5281 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     5341 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     5401 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     5461 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     5521 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     5581 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     5641 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     5701 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     5761 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     5821 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     5881 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     5941 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcc
//
