LOCUS       sequence-370760-        5839 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Testing the TEF1 promoter appended with 2 tetracycline riboswitch 
            using a green fluorescence protein as the reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5839)
  AUTHORS   Peng B, Bandari NC, Lu Z, Howard CB, Scott C, Trau M, Dumsday G, 
            Vickers CE
  TITLE     Engineering eukaryote-like regulatory circuits to expand artificial 
            control mechanisms for metabolic engineering in Saccharomyces 
            cerevisiae.
  JOURNAL   Commun Biol. 2022 Feb 16;5(1):135. doi: 10.1038/s42003-022-03070-z.
  PUBMED    35173283
REFERENCE   2  (bases 1 to 5839)
  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: "Commun 
            Biol."; date: "2022-02-16"; volume: "5(1):135. doi"; pages: " 
            10.1038/s42003-022-03070-z"
COMMENT     Sequence Label: pILGFP3A2 (1)
FEATURES             Location/Qualifiers
     source          1..5839
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             6..719
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      729..999
                     /gene="S. cerevisae PGK1"
                     /label=PGK1 terminator
                     /note="transcription terminator for 3-phosphoglycerate 
                     kinase gene"
     primer_bind     complement(1264..1280)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1264..1280)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1277..1299)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1288..1304
                     /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(1312..1318,1319..1336,1337..1342))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1357..1378
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1495..1512)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1666..2254)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1746..1765)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2425..3216,3217..3285))
                     /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     3048..3067
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3286..3390)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3458..3476
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3514..3536)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3636..3655
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3849..3871
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3863..3880
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3864..3880
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4994..5027)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        5247..5660
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
ORIGIN
        1 ccaaaatgtc taaaggtgaa gaattattca ctggtgttgt cccaattttg gttgaattag
       61 atggtgatgt taatggtcac aaattttctg tctccggtga aggtgaaggt gatgctactt
      121 acggtaaatt gaccttaaaa tttatttgta ctactggtaa attgccagtt ccatggccaa
      181 ccttagtcac tactttcggt tatggtgttc aatgttttgc gagataccca gatcatatga
      241 aacaacatga ctttttcaag tctgccatgc cagaaggtta tgttcaagaa agaactattt
      301 ttttcaaaga tgacggtaac tacaagacca gagctgaagt caagtttgaa ggtgatacct
      361 tagttaatag aatcgaatta aaaggtattg attttaaaga agatggtaac attttaggtc
      421 acaaattgga atacaactat aactctcaca atgtttacat catggctgac aaacaaaaga
      481 atggtatcaa agttaacttc aaaattagac acaacattga agatggttct gttcaattag
      541 ctgaccatta tcaacaaaat actccaattg gtgatggtcc agtcttgtta ccagacaacc
      601 attacttatc cactcaatct gccttatcca aagatccaaa cgaaaagaga gaccacatgg
      661 tcttgttaga atttgttact gctgctggta ttacccatgg tatggatgaa ttgtacaaaa
      721 gatcttaaat tgaattgaat tgaaatcgat agatcaattt ttttcttttc tctttcccca
      781 tcctttacgc taaaataata gtttatttta ttttttgaat attttttatt tatatacgta
      841 tatatagact attatttatc ttttaatgat tattaagatt tttattaaaa aaaaattcgc
      901 tcctctttta atgcctttat gcagtttttt tttcccattc gatatttcta tgttcgggtt
      961 cagcgtattt taagtttaat aactcgaaaa ttctgcgttc gttaaagctt tcgagaagga
     1021 tattatttac tagtaggact atttgcaaag ggaagggatg ctaaggtaga gggtgaacgt
     1081 tacagaaaag caggctggga agcatatttg agaagatgcg gccagcaaaa ctaaaaaact
     1141 gtattataag taaatgcatg tatactaaac tcacaaatta gagcttcaat ttaattatat
     1201 cagttattac ccgggaatct cggtcgtaat gatttaaatg catgcaagct tggcgtaatc
     1261 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     1321 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     1381 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     1441 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     1501 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     1561 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     1621 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     1681 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     1741 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     1801 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     1861 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     1921 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     1981 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     2041 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     2101 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     2161 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     2221 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     2281 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     2341 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     2401 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     2461 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     2521 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     2581 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     2641 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     2701 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     2761 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     2821 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     2881 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     2941 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     3001 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     3061 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     3121 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     3181 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     3241 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     3301 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     3361 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     3421 ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt
     3481 tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
     3541 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
     3601 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
     3661 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
     3721 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     3781 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
     3841 agggttttcc cagtcacgac gttgtaaaac gacggccagt gaattcattt aaattttttt
     3901 gattcggtaa tctccgaaca gaaggaagaa cgaaggaagg agcacagact tagattggta
     3961 tatatacgca tatgtagtgt tgaagaaaca tgaaattgcc cagtattctt aacccaactg
     4021 cacagaacaa aaacctgcag gaaacgaaga taaaacatgt ccacaaaatc atataccagt
     4081 agagctgaga ctcatgcaag tccggttgca tcgaaacttt tacgtttaat ggatgaaaag
     4141 aagaccaatt tgtgtgcttc tcttgacgtt cgttcgactg atgagctatt gaaacttgtt
     4201 gaaacgttgg gtccatacat ttgccttttg aaaacacacg ttgatatctt ggatgatttc
     4261 agttatgagg gtactgtcgt tccattgaaa gcattggcag agaaatacaa gttcttgata
     4321 tttgaggaca gaaaattcgc cgatatcggt aacacagtca aattacaata tacatcgggc
     4381 gtttaccgta tcgcagaatg gtctgatatc accaacgccc acggggttac tggtgctggt
     4441 attgttgctg gcttgaaaca aggtgcgcaa gaggtcacca aagaaccaag gggattattg
     4501 atgcttgctg aattgtcttc caagggttct ctagcacacg gtgaatatac taagggtacc
     4561 gttgatattg caaagagtga taaagatttc gttattgggt tcattgctca gaacgatatg
     4621 ggaggaagag aagaagggtt tgattggcta atcatgaccc caggtgtagg tttagacgac
     4681 aaaggcgatg cattgggtca gcagtacaga accgtcgacg aagttgtaag tggtggatca
     4741 gatatcatca ttgttggcag aggacttttc gccaagggta gagatcctaa ggttgaaggt
     4801 gaaagataca gaaatgctgg atgggaagcg taccaaaaga gaatcagcgc tccccattaa
     4861 ttatacagga aacttaatag aacaaatcac atatttaatc taatagccac ctgcattggc
     4921 acggtgcaac actacttcaa cttcatctta caaaaagatc acgtgatctg ttgtattggg
     4981 atctctagac ctaataactt cgtatagcat acattatacg aagttatatt aagggttgct
     5041 cgagaaagag ctccagaaag cgaccaccca actttggctg ataatagcgt ataaacaatg
     5101 catactttgt acgttcaaaa tacaatgcag tagatatatt tatgcatatt acatataata
     5161 catatcacat aggaagcaac aggcgcgttg gacttttaat tttcgaggac cgcgaatcct
     5221 tacatcacac ccaatccccc acaagtgatc ccccacacac catagcttca aaatgtttct
     5281 actccttttt tactcttcca gattttctcg gactccgcgc atcgccgtac cacttcaaaa
     5341 cacccaagca cagcatacta aatttcccct ctttcttcct ctagggtgtc gttaattacc
     5401 cgtactaaag gtttggaaaa gaaaaaagag accgcctcgt ttctttttct tcgtcgaaaa
     5461 aggcaataaa aatttttatc acgtttcttt ttcttgaaaa tttttttttt tgattttttt
     5521 ctctttcgat gacctcccat tgatatttaa gttaataaac ggtcttcaat ttctcaagtt
     5581 tcagtttcat ttttcttgtt ctattacaac tttttttact tcttgctcat tagaaagaaa
     5641 gcatagcaat ctaatctaag ttttaattac aaacggccta aaacatacca gatcgccacc
     5701 cgcgctttaa tctggagagg tgaagaatac gaccacctag gccaactcgc aaatcaacaa
     5761 ctctcaattc aaggcctaaa acataccaga tcgccacccg cgctttaatc tggagaggtg
     5821 aagaatacga ccacctagg
//
