LOCUS       sequence-231197-        4306 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  protein expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4306)
  TITLE     Chang lab yeast expression plasmids
REFERENCE   2  (bases 1 to 4306)
  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: Mito Carr 2
FEATURES             Location/Qualifiers
     source          1..4306
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..940
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     CDS             1975..1998
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1999..2022
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             2032..2049
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2129..2375
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        2390..2803
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        2810..2857
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2876..3250
                     /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      3316..3563
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3332..3350)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     rep_origin      complement(3638..4226)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3718..3737)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agatctaaca tccaaagacg aaaggttgaa tgaaaccttt ttgccatccg acatccacag
       61 gtccattctc acacataagt gccaaacgca acaggagggg atacactagc agcagaccgt
      121 tgcaaacgca ggacctccac tcctcttctc ctcaacaccc acttttgcca tcgaaaaacc
      181 agcccagtta ttgggcttga ttggagctcg ctcattccaa ttccttctat taggctacta
      241 acaccatgac tttattagcc tgtctatcct ggcccccctg gcgaggttca tgtttgttta
      301 tttccgaatg caacaagctc cgcattacac ccgaacatca ctccagatga gggctttctg
      361 agtgtggggt caaatagttt catgttcccc aaatggccca aaactgacag tttaaacgct
      421 gtcttggaac ctaatatgac aaaagcgtga tctcatccaa gatgaactaa gtttggttcg
      481 ttgaaatgct aacggccagt tggtcaaaaa gaaacttcca aaagtcggca taccgtttgt
      541 cttgtttggt attgattgac gaatgctcaa aaataatctc attaatgctt agcgcagtct
      601 ctctatcgct tctgaacccc ggtgcacctg tgccgaaacg caaatgggga aacacccgct
      661 ttttggatga ttatgcattg tctccacatt gtatgcttcc aagattctgg tgggaatact
      721 gctgatagcc taacgttcat gatcaaaatt taactgttct aacccctact tgacagcaat
      781 atataaacag aaggaagctg ccctgtctta aacctttttt tttatcatca ttattagctt
      841 actttcataa ttgcgactgg ttccaattga caagcttttg attttaacga cttttaacga
      901 caacttgaga agatcaaaaa acaactaatt attcgaaacg aggaattcat gagttcctca
      961 caagaatcca ccttcacctc ggcctcagcg gctgcacaag ttaatgcttc cgctttggac
     1021 ttgctcccgg tctatgccaa ggagcttatc gccggcggcg ctgctggtgc atttgcaaag
     1081 actgcggtag caccactgga gagggtcaag atcttattgc agacaagaac tgaaggtttc
     1141 cagtccctcg ggatcctcca gtccttgagg aagctgtggc aatacgaggg aattcgaggc
     1201 ttctacaaag ggaacggtgc aagcgtgctt cggattgttc catatgcagc attgcattac
     1261 atgacatatg agcagtatag gtgctggata ttgaataatt ctgcttcatc aataggtaca
     1321 gggcctgtcg ttgatctttt agctggctct gctgctggtg gtactgcagt tttgtgtaca
     1381 tacccattag acttggctag gaccaagctg gcataccagg tttcaaatgt tggccaaact
     1441 ggtaatgctt tgggaaattc tggtcaacaa caaacgtata atggcataaa agatgttttc
     1501 aagactgttt acaaggaagg aggtgcacga tctttgtatc gtggagtagg cccaactctt
     1561 attggtattc ttccatatgc tggcttgaag ttttacatat atgaagatct gaaatctcaa
     1621 gttccagacg attacaagga ttctgttata ctgaagctct cttgtggagc tttagctggt
     1681 ctatttggac aaaccctcac ttatccactg gatgttgtcc ggagacaaat gcaggttcag
     1741 agcaagcagt ctcaaaattc gagcgatggc ttccgtataa gaggaacttt tcagggtctc
     1801 ttgctcatca ttcggtgcca agggtggaga caactctttg ctggcttgag ccttaactat
     1861 gtcaaggttg tcccttcagt agctatcggt ttcacaacat acgacatgat gaaggcattg
     1921 cttggagttc ctccccgaga gagagtccat gcgtcaggtg gtaacaaggg taccttggaa
     1981 gttttgttcc agggtccaca tcatcaccac catcaccatc attgagtcga ccatcatcat
     2041 catcatcatt gagtttgtag ccttagacat gactgttcct cagttcaagt tgggcactta
     2101 cgagaagacc ggtcttgcta gattctaatc aagaggatgt cagaatgcca tttgcctgag
     2161 agatgcaggc ttcatttttg atactttttt atttgtaacc tatatagtat aggatttttt
     2221 ttgtcatttt gtttcttctc gtacgagctt gctcctgatc agcctatctc gcagctgatg
     2281 aatatcttgt ggtaggggtt tgggaaaatc attcgagttt gatgtttttc ttggtatttc
     2341 ccactcctct tcagagtaca gaagattaag tgagaccttc gtttgtgcgg atcccccaca
     2401 caccatagct tcaaaatgtt tctactcctt ttttactctt ccagattttc tcggactccg
     2461 cgcatcgccg taccacttca aaacacccaa gcacagcata ctaaatttcc cctctttctt
     2521 cctctagggt gtcgttaatt acccgtacta aaggtttgga aaagaaaaaa gagaccgcct
     2581 cgtttctttt tcttcgtcga aaaaggcaat aaaaattttt atcacgtttc tttttcttga
     2641 aaattttttt ttttgatttt tttctctttc gatgacctcc cattgatatt taagttaata
     2701 aacggtcttc aatttctcaa gtttcagttt catttttctt gttctattac aacttttttt
     2761 acttcttgct cattagaaag aaagcatagc aatctaatct aagggcggtg ttgacaatta
     2821 atcatcggca tagtatatcg gcatagtata atacgacaag gtgaggaact aaaccatggc
     2881 caagttgacc agtgccgttc cggtgctcac cgcgcgcgac gtcgccggag cggtcgagtt
     2941 ctggaccgac cggctcgggt tctcccggga cttcgtggag gacgacttcg ccggtgtggt
     3001 ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac caggtggtgc cggacaacac
     3061 cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac gccgagtggt cggaggtcgt
     3121 gtccacgaac ttccgggacg cctccgggcc ggccatgacc gagatcggcg agcagccgtg
     3181 ggggcgggag ttcgccctgc gcgacccggc cggcaactgc gtgcacttcg tggccgagga
     3241 gcaggactga cacgtccgac gcggcccgac gggtccgagg cctcggagat ccgtccccct
     3301 tttcctttgt cgatatcatg taattagtta tgtcacgctt acattcacgc cctcccccca
     3361 catccgctct aaccgaaaag gaaggagtta gacaacctga agtctaggtc cctatttatt
     3421 tttttatagt tatgttagta ttaagaacgt tatttatatt tcaaattttt cttttttttc
     3481 tgtacagacg cgtgtacgca tgtaacatta tactgaaaac cttgcttgag aaggttttgg
     3541 gacgctcgaa ggctttaatt tgcaagctgg agaccaacat gtgagcaaaa ggccagcaaa
     3601 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     3661 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     3721 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     3781 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     3841 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     3901 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     3961 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     4021 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     4081 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     4141 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     4201 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     4261 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagatc
//
