LOCUS       sequence-90267-e        4820 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cassette to replace yeast promoters with an inducible promoter 
            containing 1 lexA box.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4820)
  AUTHORS   Ottoz DS, Rudolf F, Stelling J
  TITLE     Inducible, tightly regulated and growth condition-independent 
            transcription factor in Saccharomyces cerevisiae.
  JOURNAL   Nucleic Acids Res. 2014 Jul 17. pii: gku616.
  PUBMED    25034689
REFERENCE   2  (bases 1 to 4820)
  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: "Nucleic 
            Acids Res. 2014 Jul 17. pii: gku616."
COMMENT     Sequence Label: FRP1639_PTEF-HygMX-TTEF-insul-(lexA-box)1-Pmin...
FEATURES             Location/Qualifiers
     source          1..4820
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     gene            704..2279
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        704..1047
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1048..2076
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      2082..2279
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2586..2602)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2587..2603)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2632..2652)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2632..2650)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2671..2687)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2671..2687)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2684..2706)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2695..2711
                     /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(2719..2725,2726..2743,2744..2749))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2764..2785
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2902..2919)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3073..3661)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3153..3172)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3832..4623,4624..4692))
                     /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     4455..4474
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4693..4797)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctcga
      661 tcctcgagag cttgccttgt ccccgccggg tcacccggcc agcgacatgg aggcccagaa
      721 taccctcctt gacagtcttg acgtgcgcag ctcaggggca tgatgtgact gtcgcccgta
      781 catttagccc atacatcccc atgtataatc atttgcatcc atacattttg atggccgcac
      841 ggcgcgaagc aaaaattacg gctcctcgct gcggacctgc gagcagggaa acgctcccct
      901 cacagacgcg ttgaattgtc cccacgccgc gcccctgtag agaaatataa aaggttagga
      961 tttgccactg aggttcttct ttcatatact tccttttaaa atcttgctag gatacagttc
     1021 tcacatcaca tccgaacata aacaaccatg ggtaaaaagc ctgaactcac cgcgacgtct
     1081 gtcgagaagt ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag
     1141 ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta
     1201 aatagctgcg ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc
     1261 gcgctcccga ttccggaagt gcttgacatt ggggagttca gcgagagcct gacctattgc
     1321 atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct
     1381 gttctgcagc cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg
     1441 agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc
     1501 atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc
     1561 agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa
     1621 gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc
     1681 ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc
     1741 aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag
     1801 cggaggcatc cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt
     1861 cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag
     1921 ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc
     1981 cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac
     2041 cgacgcccca gcactcgtcc gagggcaaag gaataatcag tactgacaat aaaaagattc
     2101 ttgttttcaa gaacttgtca tttgtatagt ttttttatat tgtagttgtt ctattttaat
     2161 caaatgttag cgtgatttat attttttttc gcctcgacat catctgccca gatgcgaagt
     2221 taagtgcgca gaaagtaata tcatgcgtca atcgtatgtg aatgctggtc gctatactgc
     2281 tgtcgattcg atactaacgc cgccatccag tgtttaaacg aactagtgcg gccgcaataa
     2341 tatataaacc tgtataatat aaccttgaag actatatttc ttttcgagct ccctaggtgc
     2401 tgtatatact cacagcataa ctgtatatac acccagggtc tagagcatgt gctctgtatg
     2461 tatataaaac tcttgttttc ttcttttctc ttctttcctt atacattagg acctttgcag
     2521 cataaattac tatacttcta tactagtgga tcccccgggc tgcaggaatt cgatatcaag
     2581 cttatcgata ccgtcgacct cgaggggggg cccggtaccc agcttttgtt ccctttagtg
     2641 agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     2701 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     2761 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     2821 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     2881 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     2941 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     3001 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     3061 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     3121 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     3181 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     3241 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     3301 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     3361 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     3421 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     3481 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     3541 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     3601 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     3661 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     3721 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     3781 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     3841 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     3901 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     3961 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     4021 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     4081 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     4141 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     4201 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     4261 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     4321 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     4381 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     4441 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     4501 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     4561 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     4621 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     4681 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     4741 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     4801 atttccccga aaagtgccac
//
