LOCUS       sequence-267645-        4073 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mutant form of yeast STE2 coding sequence in vector for protein 
            expression in the methanogenic yeast Pichia pastoris, with GAP 
            promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4073)
  TITLE     Wagner Lab plasmids
REFERENCE   2  (bases 1 to 4073)
  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"
COMMENT     Sequence Label: 	pGAPZa-STE2-1D4
FEATURES             Location/Qualifiers
     source          1..4073
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..477
                     /label=GAP promoter
                     /note="promoter for Pichia pastoris 
                     glyceraldehyde-3-phosphate dehydrogenase"
     CDS             join(487..543,544..741,742..753)
                     /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="MRFPSIFTAVLFAASSALAAPVNTTTEDETAQIPAEAVIGYSDLE
                     GDFDVAVLPFSNSTNNGLLFINTTIASIAAKEEGVSLEKREAEA"
     CDS             1744..1773
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1789..1806
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1886..2132
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        2147..2560
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        2567..2614
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2633..3007
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     terminator      3073..3320
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3089..3107)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     rep_origin      complement(3395..3983)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3475..3494)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tttttgtaga aatgtcttgg tgtcctcgtc caatcaggta gccatctctg aaatatctgg
       61 ctccgttgca actccgaacg acctgctggc aacgtaaaat tctccggggt aaaacttaaa
      121 tgtggagtaa tggaaccaga aacgtctctt cccttctctc tccttccacc gcccgttacc
      181 gtccctagga aattttactc tgctggagag cttcttctac ggcccccttg cagcaatgct
      241 cttcccagca ttacgttgcg ggtaaaacgg aggtcgtgta cccgacctag cagcccaggg
      301 atggaaaagt cccggccgtc gctggcaata atagcgggcg gacgcatgtc atgagattat
      361 tggaaaccac cagaatcgaa tataaaaggc gaacaccttt cccaattttg gtttctcctg
      421 acccaaagac tttaaattta atttatttgt ccctatttca atcaattgaa caactatttc
      481 gaaacgatga gatttccttc aatttttact gctgttttat tcgcagcatc ctccgcatta
      541 gctgctccag tcaacactac aacagaagat gaaacggcac aaattccggc tgaagctgtc
      601 atcggttact cagatttaga aggggatttc gatgttgctg ttttgccatt ttccaacagc
      661 acaaataacg ggttattgtt tataaatact actattgcca gcattgctgc taaagaagaa
      721 ggggtatctc tcgagaaaag agaggctgaa gctgaattca taatgtctga tgcggctcct
      781 tcattgagca atctatttta tgatccaacg tataatcctg gtcaaagcac cattaactac
      841 acttccatat atgggaatgg atctaccatc actttcgatg agttgcaagg tttagttaac
      901 agtactgtta ctcaggccat tatgtttggt gtcagatcag gcgccgctgc tttgactttg
      961 attgtcatgt ggatgacatc gagaagcaga aaaacgccga ttttcattat caaccaagtt
     1021 tcattgtttt taatcatttt gcattctgca ctctatttta aatatttact gtctaattac
     1081 tcttcagtga cttacgctct caccggattt cctcagttca tcagtagagg tgacgttcat
     1141 gtttatggtg ctacaaatat aattcaagtc cttcttgtgg cttctattga gacttcactg
     1201 gtgtttcaga taaaagttat tttcacaggc gacaacttca aaaggatagg tttgatgctg
     1261 acgtcgatat ctttcacttt agggattgct acagttacca tgtattttgt aagcgctgtt
     1321 aaaggtatga ttgtgactta taatgatgtt agtgccaccc aagataaata cttcaatgca
     1381 tccacaattt tacttgcatc ctcaataaac tttatgtcat ttgtcctggt agttaaattg
     1441 attttagcta ttagatcaag aagattcctt ggtctcaagc agttcgatag tttccatatt
     1501 ttactcataa tgtcagctca atctttgttg gtaccatcga taatattcat cctcgcatac
     1561 agtttgaaac caaaccaggg aacagatgtc ttgactactg ttgcaacatt acttgctgta
     1621 ttgtctttac cattatcatc aatgtgggcc acggctgcta ataatgcgtc gaccaccgtc
     1681 tccaagacag agaccagcca agtggcgcct gcctaaggat cctaagcggc cgccagcttt
     1741 ctagaacaaa aactcatctc agaagaggat ctgaatagcg ccgtcgacca tcatcatcat
     1801 catcattgag tttgtagcct tagacatgac tgttcctcag ttcaagttgg gcacttacga
     1861 gaagaccggt cttgctagat tctaatcaag aggatgtcag aatgccattt gcctgagaga
     1921 tgcaggcttc atttttgata cttttttatt tgtaacctat atagtatagg attttttttg
     1981 tcattttgtt tcttctcgta cgagcttgct cctgatcagc ctatctcgca gctgatgaat
     2041 atcttgtggt aggggtttgg gaaaatcatt cgagtttgat gtttttcttg gtatttccca
     2101 ctcctcttca gagtacagaa gattaagtga gaccttcgtt tgtgcggatc ccccacacac
     2161 catagcttca aaatgtttct actccttttt tactcttcca gattttctcg gactccgcgc
     2221 atcgccgtac cacttcaaaa cacccaagca cagcatacta aatttcccct ctttcttcct
     2281 ctagggtgtc gttaattacc cgtactaaag gtttggaaaa gaaaaaagag accgcctcgt
     2341 ttctttttct tcgtcgaaaa aggcaataaa aatttttatc acgtttcttt ttcttgaaaa
     2401 tttttttttt tgattttttt ctctttcgat gacctcccat tgatatttaa gttaataaac
     2461 ggtcttcaat ttctcaagtt tcagtttcat ttttcttgtt ctattacaac tttttttact
     2521 tcttgctcat tagaaagaaa gcatagcaat ctaatctaag ggcggtgttg acaattaatc
     2581 atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa ccatggccaa
     2641 gttgaccagt gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg
     2701 gaccgaccgg ctcgggttct cccgggactt cgtggaggac gacttcgccg gtgtggtccg
     2761 ggacgacgtg accctgttca tcagcgcggt ccaggaccag gtggtgccgg acaacaccct
     2821 ggcctgggtg tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc
     2881 cacgaacttc cgggacgcct ccgggccggc catgaccgag atcggcgagc agccgtgggg
     2941 gcgggagttc gccctgcgcg acccggccgg caactgcgtg cacttcgtgg ccgaggagca
     3001 ggactgacac gtccgacgcg gcccgacggg tccgaggcct cggagatccg tccccctttt
     3061 cctttgtcga tatcatgtaa ttagttatgt cacgcttaca ttcacgccct ccccccacat
     3121 ccgctctaac cgaaaaggaa ggagttagac aacctgaagt ctaggtccct atttattttt
     3181 ttatagttat gttagtatta agaacgttat ttatatttca aatttttctt ttttttctgt
     3241 acagacgcgt gtacgcatgt aacattatac tgaaaacctt gcttgagaag gttttgggac
     3301 gctcgaaggc tttaatttgc aagctggaga ccaacatgtg agcaaaaggc cagcaaaagg
     3361 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     3421 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     3481 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     3541 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     3601 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     3661 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     3721 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     3781 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     3841 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     3901 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     3961 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     4021 agtggaacga aaactcacgt taagggattt tggtcatgca tgagatcaga tct
//
