LOCUS       sequence-159456-        6454 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pheromone mediated alpha pheromone expression in yeast from the 
            mfalpha 1 gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6454)
  AUTHORS   Williams TC, Nielsen LK, Vickers CE
  TITLE     Engineered quorum sensing using pheromone-mediated cell-to-cell 
            communication in Saccharomyces cerevisiae.
  JOURNAL   ACS Synth Biol. 2013 Mar 15;2(3):136-49. doi: 10.1021/sb300110b. 
            Epub 2013 Jan 7.
  PUBMED    23656437
REFERENCE   2  (bases 1 to 6454)
  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: "10.1021/sb300110b";
            journalName: "ACS Synth Biol"; date: "2013-03-15- 15"; volume: "2"; 
            issue: "3"; pages: "136-49"
COMMENT     Sequence Label: pTCW005
FEATURES             Location/Qualifiers
     source          1..6454
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        317..503
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             504..1163
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL
                     ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(1423..1878)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1560..1581)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1772..1791
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2008..2030
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2022..2039
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2023..2039
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2049..2068
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2049..2067
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2828..2884,2885..3082,3083..3094)
                     /codon_start=1
                     /gene="MF-alpha-1"
                     /product="N-terminal secretion signal from S. cerevisiae 
                     alpha-factor"
                     /label=alpha-factor secretion signal
                     /note="Cleavage by the Kex2 protease occurs after the 
                     dibasic KR sequence. The EA dipeptides are then removed by 
                     dipeptidyl aminopeptidase A."
                     /translation="MRFPSIFTAVLFAASSALAAPVNTTTEDETAQIPAEAVIGYLDLE
                     GDFDVAVLPFSNSTNNGLLFINTTIASIAAKEEGVSLDKREAEA"
     primer_bind     complement(3627..3643)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3627..3643)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3680..3700)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3680..3698)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3719..3735)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3719..3735)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3732..3754)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3743..3759
                     /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(3767..3773,3774..3791,3792..3797))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(3950..3967)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4121..4709)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4201..4220)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4880..5671,5672..5740))
                     /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     5503..5522
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5741..5845)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    5882..6385
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     6427..6445
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgcgtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataaatt cccgttttaa gagcttggtg agcgctagga gtcactgcca ggtatcgttt
      241 gaacacggca ttagtcaggg aagtcataac acagtccttt cccgcaattt tctttttcta
      301 ttactcttgg cctcctctag tacactctat atttttttat gcctcggtaa tgattttcat
      361 tttttttttt cccctagcgg atgactcttt ttttttctta gcgattggca ttatcacata
      421 atgaattata cattatataa agtaatgtga tttcttcgaa gaatatacta aaaaatgagc
      481 aggcaagata aacgaaggca aagatgacag agcagaaagc cctagtaaag cgtattacaa
      541 atgaaaccaa gattcagatt gcgatctctt taaagggtgg tcccctagcg atagagcact
      601 cgatcttccc agaaaaagag gcagaagcag tagcagaaca ggccacacaa tcgcaagtga
      661 ttaacgtcca cacaggtata gggtttctgg accatatgat acatgctctg gccaagcatt
      721 ccggctggtc gctaatcgtt gagtgcattg gtgacttaca catagacgac catcacacca
      781 ctgaagactg cgggattgct ctcggtcaag cttttaaaga ggccctactg gcgcgtggag
      841 taaaaaggtt tggatcagga tttgcgcctt tggatgaggc actttccaga gcggtggtag
      901 atctttcgaa caggccgtac gcagttgtcg aacttggttt gcaaagggag aaagtaggag
      961 atctctcttg cgagatgatc ccgcattttc ttgaaagctt tgcagaggct agcagaatta
     1021 ccctccacgt tgattgtctg cgaggcaaga atgatcatca ccgtagtgag agtgcgttca
     1081 aggctcttgc ggttgccata agagaagcca cctcgcccaa tggtaccaac gatgttccct
     1141 ccaccaaagg tgttcttatg tagtgacacc gattatttaa agctgcagca tacgatatat
     1201 atacatgtgt atatatgtat acctatgaat gtcagtaagt atgtatacga acagtatgat
     1261 actgaagatg acaaggtaat gcatcattct atacgtgtca ttctgaacga ggcgcgcttt
     1321 ccttttttct ttttgctttt tctttttttt tctcttgaac tcgacggatc tatgcggtgt
     1381 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggaaattgta aacgttaata
     1441 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     1501 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     1561 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     1621 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     1681 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     1741 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     1801 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     1861 cgccgctaca gggcgcgtcg cgccattcgc cattcaggct gcgcaactgt tgggaagggc
     1921 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc
     1981 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg
     2041 agcgcgcgta atacgactca ctatagggcg aattgggtac cgggcccccc ctcgagatca
     2101 acaacagggt cagcagcagc aacaaggcca ctccagttca ttctcagctt tggcttccat
     2161 ggcaagttcc tacctgggca ataactccaa ttcaaattcg agttatgggg gccagcaaca
     2221 ggctaatgag tatggtagac cgcaacagaa tggtcaacag caatccaatg agtacggaag
     2281 accgcaatac ggcggaaacc agaactccaa tggacagcac gaatccttca atttttctgg
     2341 caacttttct caacagaaca ataacggcaa ccagaaccgc tactgaacga tgattcagtt
     2401 cgccttctat cctttgttta cgtatttgtt tatatatata actttatttt tttttattaa
     2461 ttgggctgca agacaatttt gttgtcagtg atgcctcaat ccttcttttg cttccatatt
     2521 taccatgtgg accctttcaa aacagagttg tatctctgca ggatgccctt tttgacgtat
     2581 tgaatggcat aattgcactg tcacttttcg cgctgtctca ttttggtgcg atgatgaaac
     2641 aaacatgaaa cgtctgtaat ttgaaacaaa taacgtaatt ctcgggattg gttttattta
     2701 aatgacaatg taagagtggc tttgtaaggt atgtgttgct cttaaaatat ttggatacga
     2761 catcctttat cttttttcct ttaagagcag gatataagcc atcaagtttc tgaaaatcaa
     2821 agaattcatg agatttcctt caatttttac tgcagtttta ttcgcagcat cctccgcatt
     2881 agctgctcca gtcaacacta caacagaaga tgaaacggca caaattccgg ctgaagctgt
     2941 catcggttac ttagatttag aaggggattt cgatgttgct gttttgccat tttccaacag
     3001 cacaaataac gggttattgt ttataaatac tactattgcc agcattgctg ctaaagaaga
     3061 aggggtatct ttggataaaa gagaggctga agcttggcat tggttgcaac taaaacctgg
     3121 ccaaccaatg tacaagagag aagccgaagc tgaagcttgg cattggctgc aactaaagcc
     3181 tggccaacca atgtacaaaa gagaagccga cgctgaagct tggcattggc tgcaactaaa
     3241 gcctggccaa ccaatgtaca aaagagaagc cgacgctgaa gcttggcatt ggttgcagtt
     3301 aaaacccggc caaccaatgt actaagcccg actgataaca acagtgtaga tgtaacaaag
     3361 tcgactttgt tcccactgta cttttagctc gtacaaaata caatatactt ttcatttctc
     3421 cgtaaacaac atgttttccc atgtaatatc cttttctatt tttcgttccg ttaccaactt
     3481 tacacatact ttatatagct attcacttct atacactaaa aaactaagac aattttaatt
     3541 ttgctgcctg ccatatttca atttgttata aattcctata atttatccta ttagtagcta
     3601 aaaaaagatg aatgtgaatc gaatcggatc cactagttct agagcggccg ccaccgcggt
     3661 ggagctccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt
     3721 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac ataggagccg
     3781 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag gtaactcaca ttaattgcgt
     3841 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     3901 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     3961 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4021 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4081 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4141 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4201 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4261 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     4321 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     4381 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     4441 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     4501 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     4561 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     4621 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     4681 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     4741 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     4801 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     4861 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     4921 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     4981 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     5041 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     5101 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     5161 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     5221 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     5281 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     5341 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     5401 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     5461 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     5521 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     5581 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     5641 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     5701 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     5761 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     5821 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gggtcctttt
     5881 catcacgtgc tataaaaata attataattt aaatttttta atataaatat ataaattaaa
     5941 aatagaaagt aaaaaaagaa attaaagaaa aaatagtttt tgttttccga agatgtaaaa
     6001 gactctaggg ggatcgccaa caaatactac cttttatctt gctcttcctg ctctcaggta
     6061 ttaatgccga attgtttcat cttgtctgtg tagaagacca cacacgaaaa tcctgtgatt
     6121 ttacatttta cttatcgtta atcgaatgta tatctattta atctgctttt cttgtctaat
     6181 aaatatatat gtaaagtacg ctttttgttg aaatttttta aacctttgtt tatttttttt
     6241 tcttcattcc gtaactcttc taccttcttt atttactttc taaaatccaa atacaaaaca
     6301 taaaaataaa taaacacaga gtaaattccc aaattattcc atcattaaaa gatacgaggc
     6361 gcgtgtaagt tacaggcaag cgatccgtcc taagaaacca ttattatcat gacattaacc
     6421 tataaaaata ggcgtatcac gaggcccttt cgtc
//
