LOCUS       sequence-214068-        4573 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human gamma-actin (codon is optimised for expression in 
            Pichia pastoris) fused with thymosin beta and a His tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4573)
  AUTHORS   Hatano T, Alioto S, Roscioli E, Palani S, Clarke ST, Kamnev A, 
            Hernandez-Fernaud JR, Sivashanmugam L, Chapa-Y-Lazo B, Jones AME, 
            Robinson RC, Sampath K, Mishima M, McAinsh AD, Goode BL, 
            Balasubramanian MK
  TITLE     Rapid production of pure recombinant actin isoforms in Pichia 
            pastoris.
  JOURNAL   J Cell Sci. 2018 Mar 13. pii: jcs.213827. doi: 10.1242/jcs.213827.
  PUBMED    29535210
REFERENCE   2  (bases 1 to 4573)
  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 Cell Sci.
            2018 Mar 13. pii: jcs.213827. doi: 10.1242/jcs.213827."
COMMENT     Sequence Label: pPICZc-HsACTG1*-thymosinB-8His
FEATURES             Location/Qualifiers
     source          1..4573
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..940
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     regulatory      949..958
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2257..2277
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2281..2304
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     terminator      2396..2642
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        2657..3070
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        3077..3124
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3143..3517
                     /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      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 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 aggaattcgc caccatggaa
      961 gaagagatcg ccgctttggt tatcgacaac ggttctggta tgtgtaaggc tggtttcgct
     1021 ggtgatgatg ctccaagagc tgttttccca tccatcgttg gtagaccaag acaccagggt
     1081 gttatggttg gtatgggtca aaaggactcc tacgttggtg acgaagctca atccaagaga
     1141 ggtatcttga ccctgaagta cccaatcgag cacggtattg tcactaactg ggacgacatg
     1201 gaaaagatct ggcaccacac tttctacaac gagttgagag ttgctccaga ggaacaccca
     1261 gttttgttga ctgaagctcc attgaaccca aaggccaaca gagaaaagat gacccagatc
     1321 atgttcgaga ctttcaacac cccagctatg tacgttgcta tccaggctgt tttgtccttg
     1381 tacgcttccg gtagaactac cggtatcgtt atggattctg gtgacggtgt tactcacacc
     1441 gttccaatct acgaaggtta cgctttgcca cacgccatct tgagattgga tttggctggt
     1501 agagacttga ccgactacct gatgaagatc ttgaccgaga gaggttactc cttcactact
     1561 actgctgaga gagagatcgt cagggacatc aaagaaaagt tgtgctacgt cgccctggac
     1621 ttcgaacaag aaatggctac tgctgcttcc tcatcctctt tggaaaagtc ctacgaattg
     1681 ccagacggtc aggttatcac tatcggtaac gagagattca gatgcccaga ggctttgttc
     1741 caaccatcct tccttggtat ggaatcctgt ggtatccacg agactacctt caactccatc
     1801 atgaagtgtg acgtcgacat cagaaaggac ttgtacgcca acaccgtttt gtccggtggt
     1861 actactatgt acccaggtat cgctgacaga atgcagaaag agattactgc tttggcccca
     1921 tccaccatga agatcaagat tattgctcca ccagagagga agtactccgt ttggattggt
     1981 ggttctatct tggcttcctt gtccaccttc cagcagatgt ggatttccaa gcaagaatac
     2041 gacgaatccg gtccttccat cgttcacaga aagtgttttg ctagccgtgg cggcagcggt
     2101 gggagctctg gcggtagcgc ttccgataag ccagatatgg cggaaatcga aaagttcgat
     2161 aaatctaaac tgaagaaaac tgagactcaa gaaaagaacc cgctgccgag caaggaaacc
     2221 atcgagcagg agaagcaggc gggtgaatcc ggtaccgaaa atttatattt tcaagggccc
     2281 caccaccacc accaccacca ccacgaagaa taacttaagt caattcgcct tagacatgac
     2341 tgttcctcag ttcaagttgg gcacttacga gaagaccggt cttgctagat tctaatcaag
     2401 aggatgtcag aatgccattt gcctgagaga tgcaggcttc atttttgata cttttttatt
     2461 tgtaacctat atagtatagg attttttttg tcattttgtt tcttctcgta cgagcttgct
     2521 cctgatcagc ctatctcgca gctgatgaat atcttgtggt aggggtttgg gaaaatcatt
     2581 cgagtttgat gtttttcttg gtatttccca ctcctcttca gagtacagaa gattaagtga
     2641 gaccttcgtt tgtgcggatc ccccacacac catagcttca aaatgtttct actccttttt
     2701 tactcttcca gattttctcg gactccgcgc atcgccgtac cacttcaaaa cacccaagca
     2761 cagcatacta aatttcccct ctttcttcct ctagggtgtc gttaattacc cgtactaaag
     2821 gtttggaaaa gaaaaaagag accgcctcgt ttctttttct tcgtcgaaaa aggcaataaa
     2881 aatttttatc acgtttcttt ttcttgaaaa tttttttttt tgattttttt ctctttcgat
     2941 gacctcccat tgatatttaa gttaataaac ggtcttcaat ttctcaagtt tcagtttcat
     3001 ttttcttgtt ctattacaac tttttttact tcttgctcat tagaaagaaa gcatagcaat
     3061 ctaatctaag ggcggtgttg acaattaatc atcggcatag tatatcggca tagtataata
     3121 cgacaaggtg aggaactaaa ccatggccaa gttgaccagt gccgttccgg tgctcaccgc
     3181 gcgcgacgtc gccggagcgg tcgagttctg gaccgaccgg ctcgggttct cccgggactt
     3241 cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg accctgttca tcagcgcggt
     3301 ccaggaccag gtggtgccgg acaacaccct ggcctgggtg tgggtgcgcg gcctggacga
     3361 gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc cgggacgcct ccgggccggc
     3421 catgaccgag atcggcgagc agccgtgggg gcgggagttc gccctgcgcg acccggccgg
     3481 caactgcgtg 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 atc
//
