LOCUS       sequence-251755-        4328 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Saporin L3 in pichia.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4328)
  AUTHORS   Yuan H, Du Q, Sturm MB, Schramm VL
  TITLE     Soapwort Saporin L3 Expression in Yeast, Mutagenesis, and RNA 
            Substrate Specificity.
  JOURNAL   Biochemistry. 2015 Jul 28;54(29):4565-74. doi: 
            10.1021/acs.biochem.5b00405. Epub 2015 Jul 14.
  PUBMED    26091305
REFERENCE   2  (bases 1 to 4328)
  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/acs.biochem.5b00405"; journalName: "Biochemistry"; date: 
            "2015-07-28- 28"; volume: "54"; issue: "29"; pages: "4565-74"
COMMENT     Sequence Label: pPICZalphaB-SapL3
FEATURES             Location/Qualifiers
     source          1..4328
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..939
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     CDS             join(940..996,997..1194,1195..1206)
                     /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             2008..2037
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2053..2070
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2150..2396
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        2411..2824
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        2831..2878
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2897..3271
                     /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      3337..3584
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3353..3371)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     rep_origin      complement(3659..4247)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3739..3758)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gatctaacat ccaaagacga aaggttgaat gaaacctttt tgccatccga catccacagg
       61 tccattctca cacataagtg ccaaacgcaa caggagggga tacactagca gcagaccgtt
      121 gcaaacgcag gacctccact cctcttctcc tcaacaccca cttttgccat cgaaaaacca
      181 gcccagttat tgggcttgat tggagctcgc tcattccaat tccttctatt aggctactaa
      241 caccatgact ttattagcct gtctatcctg gcccccctgg cgaggttcat gtttgtttat
      301 ttccgaatgc aacaagctcc gcattacacc cgaacatcac tccagatgag ggctttctga
      361 gtgtggggtc aaatagtttc atgttcccca aatggcccaa aactgacagt ttaaacgctg
      421 tcttggaacc taatatgaca aaagcgtgat ctcatccaag atgaactaag tttggttcgt
      481 tgaaatgcta acggccagtt ggtcaaaaag aaacttccaa aagtcggcat accgtttgtc
      541 ttgtttggta ttgattgacg aatgctcaaa aataatctca ttaatgctta gcgcagtctc
      601 tctatcgctt ctgaaccccg gtgcacctgt gccgaaacgc aaatggggaa acacccgctt
      661 tttggatgat tatgcattgt ctccacattg tatgcttcca agattctggt gggaatactg
      721 ctgatagcct aacgttcatg atcaaaattt aactgttcta acccctactt gacagcaata
      781 tataaacaga aggaagctgc cctgtcttaa accttttttt ttatcatcat tattagctta
      841 ctttcataat tgcgactggt tccaattgac aagcttttga ttttaacgac ttttaacgac
      901 aacttgagaa gatcaaaaaa caactaatta ttcgaaacga tgagatttcc ttcaattttt
      961 actgctgttt tattcgcagc atcctccgca ttagctgctc cagtcaacac tacaacagaa
     1021 gatgaaacgg cacaaattcc ggctgaagct gtcatcggtt actcagattt agaaggggat
     1081 ttcgatgttg ctgttttgcc attttccaac agcacaaata acgggttatt gtttataaat
     1141 actactattg ccagcattgc tgctaaagaa gaaggggtat ctctcgagaa aagagaggct
     1201 gaagctgcag gcgtaattat atatgaattg aatctccaag gtactaccaa ggcgcaatac
     1261 tcgacatttc tcaaacagct ccgtgacgat ataaaggatc caaacctcca ttacggtggt
     1321 accaatctac ccgtgataaa aagacctgtg ggccccccga aattccttag agttaatctc
     1381 aaagcttcta ctggaactgt ctcacttgcc gtacagcgta gtaacttgta tgtggccgcg
     1441 tatcttgcta agaataacaa caaacaattt cgggcatatt actttaaggg cttccaaatt
     1501 actacgaatc agttaaacaa ccttttccca gaggcaactg gtgtctcaaa tcagcaagaa
     1561 ttagggtatg gtgaatcata tccgcaaata caaaacgcag ccggggtaac aagacagcaa
     1621 gcgggcttgg ggatcaagaa acttgctgag tccatgacaa aagtgaacgg ggttgcgcgt
     1681 gttgaaaaag atgaagcact gtttttgttg atcgttgttc aaatggtggg tgaagcagcg
     1741 cgatttaagt acatagagaa tttggtactt aacaattttg atacagcaaa ggaggttgaa
     1801 cccgttcctg atcgtgtgat tatattggaa aataattggg gtctgctttc aagagctgcc
     1861 aaaactgcaa acaatggagt gttccaaact ccgcttgttt tgacaagtta tgctgtccct
     1921 ggcgtcgagt ggagggtgac tactgtggcg gaagtggaaa tagggatctt tctgaacgta
     1981 gacaacaatg cggccgccag ctttctagaa caaaaactca tctcagaaga ggatctgaat
     2041 agcgccgtcg accatcatca tcatcatcat tgagtttgta gccttagaca tgactgttcc
     2101 tcagttcaag ttgggcactt acgagaagac cggtcttgct agattctaat caagaggatg
     2161 tcagaatgcc atttgcctga gagatgcagg cttcattttt gatacttttt tatttgtaac
     2221 ctatatagta taggattttt tttgtcattt tgtttcttct cgtacgagct tgctcctgat
     2281 cagcctatct cgcagctgat gaatatcttg tggtaggggt ttgggaaaat cattcgagtt
     2341 tgatgttttt cttggtattt cccactcctc ttcagagtac agaagattaa gtgagacctt
     2401 cgtttgtgcg gatcccccac acaccatagc ttcaaaatgt ttctactcct tttttactct
     2461 tccagatttt ctcggactcc gcgcatcgcc gtaccacttc aaaacaccca agcacagcat
     2521 actaaatttc ccctctttct tcctctaggg tgtcgttaat tacccgtact aaaggtttgg
     2581 aaaagaaaaa agagaccgcc tcgtttcttt ttcttcgtcg aaaaaggcaa taaaaatttt
     2641 tatcacgttt ctttttcttg aaaatttttt tttttgattt ttttctcttt cgatgacctc
     2701 ccattgatat ttaagttaat aaacggtctt caatttctca agtttcagtt tcatttttct
     2761 tgttctatta caactttttt tacttcttgc tcattagaaa gaaagcatag caatctaatc
     2821 taagggcggt gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa
     2881 ggtgaggaac taaaccatgg ccaagttgac cagtgccgtt ccggtgctca ccgcgcgcga
     2941 cgtcgccgga gcggtcgagt tctggaccga ccggctcggg ttctcccggg acttcgtgga
     3001 ggacgacttc gccggtgtgg tccgggacga cgtgaccctg ttcatcagcg cggtccagga
     3061 ccaggtggtg ccggacaaca ccctggcctg ggtgtgggtg cgcggcctgg acgagctgta
     3121 cgccgagtgg tcggaggtcg tgtccacgaa cttccgggac gcctccgggc cggccatgac
     3181 cgagatcggc gagcagccgt gggggcggga gttcgccctg cgcgacccgg ccggcaactg
     3241 cgtgcacttc gtggccgagg agcaggactg acacgtccga cgcggcccga cgggtccgag
     3301 gcctcggaga tccgtccccc ttttcctttg tcgatatcat gtaattagtt atgtcacgct
     3361 tacattcacg ccctcccccc acatccgctc taaccgaaaa ggaaggagtt agacaacctg
     3421 aagtctaggt ccctatttat ttttttatag ttatgttagt attaagaacg ttatttatat
     3481 ttcaaatttt tctttttttt ctgtacagac gcgtgtacgc atgtaacatt atactgaaaa
     3541 ccttgcttga gaaggttttg ggacgctcga aggctttaat ttgcaagctg gagaccaaca
     3601 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     3661 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     3721 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     3781 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     3841 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     3901 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     3961 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4021 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4081 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4141 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4201 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4261 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     4321 tgagatca
//
