LOCUS       sequence-252827-        4574 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses E coli Bir A protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4574)
  AUTHORS   de la Cruz S, Alcocer M, Madrid R, Garcia A, Martin R, Gonzalez I, 
            Garcia T
  TITLE     Production of in vivo biotinylated scFv specific to almond (Prunus 
            dulcis) proteins by recombinant Pichia pastoris.
  JOURNAL   J Biotechnol. 2016 Jun 10;227:112-9. doi: 
            10.1016/j.jbiotec.2016.04.024. Epub 2016 Apr 13.
  PUBMED    27085890
REFERENCE   2  (bases 1 to 4574)
  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: "J 
            Biotechnol."; date: "2016-06-10"; volume: "227:112-9. doi"; pages: "
            10.1016/j.jbiotec.2016.04.024. Epub 2016 Apr 13"
COMMENT     Sequence Label: pMJA180
FEATURES             Location/Qualifiers
     source          1..4574
                     /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             1213..2178
                     /codon_start=1
                     /gene="birA"
                     /product="E. coli biotin protein ligase"
                     /label=BirA
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             1213..2175
                     /codon_start=1
                     /product="E. coli biotin protein ligase"
                     /label=BirA
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             2202..2231
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2247..2264
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2344..2590
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        2605..3018
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        3025..3072
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3091..3489
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     terminator      3583..3830
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3599..3617)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     rep_origin      complement(3905..4493)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3985..4004)
                     /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 gaagctgaat tcatgaagga taacaccgtg ccactgaaat tgattgccct gttagcgaac
     1261 ggtgaatttc actctggcga gcagttgggt gaaacgctgg gaatgagccg ggcggctatt
     1321 aataaacaca ttcagacact gcgtgactgg ggcgttgatg tctttaccgt tccgggtaaa
     1381 ggatacagcc tgcctgagcc tatccagtta cttaatgcta aacagatatt gggtcagctg
     1441 gatggcggta gtgtagccgt gctgccagtg attgactcca cgaatcagta ccttcttgat
     1501 cgtatcggag agcttaaatc gggcgatgct tgcattgcag aataccagca ggctggccgt
     1561 ggtcgccggg gtcggaaatg gttttcgcct tttggcgcaa acttatattt gtcgatgttc
     1621 tggcgtctgg aacaaggccc ggcggcggcg attggtttaa gtctggttat cggtatcgtg
     1681 atggcggaag tattacgcaa gctgggtgca gataaagttc gtgttaaatg gcctaatgac
     1741 ctctatctgc aggatcgcaa gctggcaggc attctggtgg agctgactgg caaaactggc
     1801 gatgcggcgc aaatagtcat tggagccggg atcaacatgg caatgcgccg tgttgaagag
     1861 agtgtcgtta atcaggggtg gatcacgctg caggaagcgg ggatcaatct cgatcgtaat
     1921 acgttggcgg ccatgctaat acgtgaatta cgtgctgcgt tggaactctt cgaacaagaa
     1981 ggattggcac cttatctgtc gcgctgggaa aagctggata attttattaa tcgcccagtg
     2041 aaacttatca ttggtgataa agaaatattt ggcatttcac gcggaataga caaacagggg
     2101 gctttattac ttgagcagga tggaataata aaaccctgga tgggcggtga aatatccctg
     2161 cgtagtgcag aaaaataacc gcggcggccg ccagctttct agaacaaaaa ctcatctcag
     2221 aagaggatct gaatagcgcc gtcgaccatc atcatcatca tcattgagtt tgtagcctta
     2281 gacatgactg ttcctcagtt caagttgggc acttacgaga agaccggtct tgctagattc
     2341 taatcaagag gatgtcagaa tgccatttgc ctgagagatg caggcttcat ttttgatact
     2401 tttttatttg taacctatat agtataggat tttttttgtc attttgtttc ttctcgtacg
     2461 agcttgctcc tgatcagcct atctcgcagc tgatgaatat cttgtggtag gggtttggga
     2521 aaatcattcg agtttgatgt ttttcttggt atttcccact cctcttcaga gtacagaaga
     2581 ttaagtgaga ccttcgtttg tgcggatccc ccacacacca tagcttcaaa atgtttctac
     2641 tcctttttta ctcttccaga ttttctcgga ctccgcgcat cgccgtacca cttcaaaaca
     2701 cccaagcaca gcatactaaa tttcccctct ttcttcctct agggtgtcgt taattacccg
     2761 tactaaaggt ttggaaaaga aaaaagagac cgcctcgttt ctttttcttc gtcgaaaaag
     2821 gcaataaaaa tttttatcac gtttcttttt cttgaaaatt tttttttttg atttttttct
     2881 ctttcgatga cctcccattg atatttaagt taataaacgg tcttcaattt ctcaagtttc
     2941 agtttcattt ttcttgttct attacaactt tttttacttc ttgctcatta gaaagaaagc
     3001 atagcaatct aatctaaggg cggtgttgac aattaatcat cggcatagta tatcggcata
     3061 gtataatacg acaaggtgag gaactaaacc atggccaagc ctttgtctca agaagaatcc
     3121 accctcattg aaagagcaac ggctacaatc aacagcatcc ccatctctga agactacagc
     3181 gtcgccagcg cagctctctc tagcgacggc cgcatcttca ctggtgtcaa tgtatatcat
     3241 tttactgggg gaccttgtgc agaactcgtg gtgctgggca ctgctgctgc tgcggcagct
     3301 ggcaacctga cttgtatcgt cgcgatcgga aatgagaaca ggggcatctt gagcccctgc
     3361 ggacggtgcc gacaggtgct tctcgatctg catcctggga tcaaagccat agtgaaggac
     3421 agtgatggac agccgacggc agttgggatt cgtgaattgc tgccctctgg ttatgtgtgg
     3481 gagggctaag cacttcgtgg ccgaggagca ggactgacac gtccgacgcg gcccgacggg
     3541 tccgaggcct cggagatccg tccccctttt cctttgtcga tatcatgtaa ttagttatgt
     3601 cacgcttaca ttcacgccct ccccccacat ccgctctaac cgaaaaggaa ggagttagac
     3661 aacctgaagt ctaggtccct atttattttt ttatagttat gttagtatta agaacgttat
     3721 ttatatttca aatttttctt ttttttctgt acagacgcgt gtacgcatgt aacattatac
     3781 tgaaaacctt gcttgagaag gttttgggac gctcgaaggc tttaatttgc aagctggaga
     3841 ccaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     3901 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     3961 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4021 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4081 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4141 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4201 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4261 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4321 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     4381 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     4441 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     4501 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     4561 tggtcatgag atca
//
