LOCUS       sequence-358139-        3208 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  HiFi-compatible destination vector for secreted overexpression of 
            thrombin-cleavable, C-terminal Protein A-10His-TwinStrep tagged 
            proteins with ExpreS2 Platform.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3208)
  AUTHORS   Coker JA, Katis VL, Fairhead M, Schwenzer A, Clemmensen SB, Frandsen
            BU, de Jongh WA, Gileadi O, Burgess-Brown NA, Marsden BD, Midwood 
            KS, Yue WW
  TITLE     FAS2FURIOUS: Moderate-Throughput Secreted Expression of Difficult 
            Recombinant Proteins in Drosophila S2 Cells.
  JOURNAL   Front Bioeng Biotechnol. 2022 May 5;10:871933. doi: 
            10.3389/fbioe.2022.871933. eCollection 2022.
  PUBMED    35600892
REFERENCE   2  (bases 1 to 3208)
  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: "Front 
            Bioeng Biotechnol."; date: "2022-05-5"; volume: "10:871933. doi"; 
            pages: " 10.3389/fbioe.2022.871933. eCollection 2022"
COMMENT     Sequence Label: pExpreS2-1-CR-PA-10H-SIII
FEATURES             Location/Qualifiers
     source          1..3208
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    47..168
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(84..103)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     138..157
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     intron          881..1013
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     sig_peptide     1036..1089
                     /codon_start=1
                     /product="Drosophila BiP signal sequence"
                     /label=BiP signal sequence
                     /translation="MKLCILLAVVAFVGLSLG"
     CDS             1096..1113
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1513..1542
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             join(1552..1575,1576..1611,1612..1635)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     polyA_signal    1680..1809
                     /gene="immediate-early gene 2 from the Orgyia pseudotsugata
                     multicapsid nucleopolyhedrovirus"
                     /label=OpIE-2 poly(A) signal
                     /note="baculovirus polyadenylation signal"
     rep_origin      complement(1944..2532)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2024..2043)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2789..2807
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2811..2820
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2817..3191
                     /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"
ORIGIN
        1 cgccggtccg acggcggccc acgggtccca ggggggtcga cctcgaaact tgtttattgc
       61 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
      121 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtcttcac
      181 gtaataagtg tgcggctagc agtcaactag tccaaactca ttaatgtatc ttatcatgtc
      241 ttcacgtaat aagtgtgcgg ctagcagtca actactagtg aatgccctac tagaagatgt
      301 gtgttgcaca aaatgtccct ggaataacca atttgaagtg cagatagcag taaacgtaag
      361 ctaatatgaa tattatttaa ctgtaatgtt ttaatatcgc tggacattac taataaaccc
      421 actataaaca catgtacata tgtatgtttt ggcatacaat gagtagttgg ggaaaaaatg
      481 tgtaaaagca ccgtgaccat cacagcataa agataaccag ctgaagtatc gaatatgagt
      541 aacccccaaa ttgaatcaca tgccgcaact gataggaccc atggaagtac actcttcatg
      601 gcgatataca agacacacac aagcacgaac acccagttgc ggaggaaatt ctccgtaaat
      661 gaaaacccaa tcggcgaaca attcataccc atatatggta aaagttttga acgcgagcga
      721 acttaagagc gccggagtat aaatagaggc gcttcgtcta cggagcgaca attcaattca
      781 aacaagcaaa gtgaacacgt cgctaagcga aagctaagca aataaacaag cgcagctgaa
      841 caagctaaac aatctgcaga agcttggtac cctcgagctc gtaagtatca aggttacaag
      901 acaggtttaa ggagaccaat agaaactggg cttgtcgaga cagagaagac tcttgcgttt
      961 ctgataggca cctattggtc ttactgacat ccactttgcc tttctctcca caggaattct
     1021 ggatccgatc tcaatatgaa gttatgcata ttactggccg tcgtggcctt tgttggcctc
     1081 tcgctcggga gatctctggt gccacgcgga tcggccgccc agcatgacga ggccgtggat
     1141 aacaagttca ataaggagca gcagaacgcc ttctacgaga tcctgcacct gcccaacctg
     1201 aatgaggagc agcgcaatgc cttcattcag agcctgaagg acgatccatc ccagtcggcc
     1261 aacctgctgg ccgaggccaa gaagctgaat gacgcccagg ccccgaaggt ggacaacaag
     1321 tttaataagg agcagcagaa tgccttttat gagatcctgc atctgccaaa tctgaacgag
     1381 gagcagcgca acgcctttat ccagtccctg aaggatgatc caagccagtc cgccaattta
     1441 ttagctgaag ccaagaagct gaacgatgcc caggccccca aggtggatgc caactcctcg
     1501 agcgtgccat cgcaccatca tcaccatcac catcaccacc atggttctgg ttggagtcac
     1561 ccacaattcg agaaaggtgg tggctctggt ggcggttcag gcggctccgc ctggtcccat
     1621 ccacagtttg aaaagtgaac cggtcatatg cggccgcgga tcgatcgata tctgactaaa
     1681 tcttagtttg tattgtcatg ttttaataca atatgttatg tttaaatatg tttttaataa
     1741 attttataaa ataatttcaa cttttattgt aacaacattg tccatttaca cactcctttc
     1801 aagcgcgtgg gactcgatgc tcggcgccac tcaaaggcgg taatacggtt atccacagaa
     1861 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     1921 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     1981 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2041 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2101 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     2161 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     2221 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     2281 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     2341 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     2401 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     2461 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     2521 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     2581 aactcacgtt aagggatttt gcatgcgcta agcgggcttt ataaaacggg ctgcgggacc
     2641 agttttcata tcactaccgt ttgagttctt gtgctgtgtg gatactcctc ccgacaccga
     2701 attaattcgg atctctgcaa gggattttgg tcatgaacaa taaaactgtc tgcttacata
     2761 aacagtaata caaggggtgt tcatagtata atacgactca ctataggagg gccaccatgg
     2821 ccaagttgac cagtgccgtt ccggtgctca ccgcgcgcga cgtcgccgga gcggtcgagt
     2881 tctggaccga ccggctcggg ttcagccggg acttcgtgga ggacgacttc gccggtgtgg
     2941 tccgggacga cgtgaccctg ttcatcagcg cggtccagga ccaggtggtg ccggacaaca
     3001 ccctggcctg ggtgtgggtg cgcggcctgg acgagctgta cgccgagtgg tcggaggtcg
     3061 tgtccacgaa cttccgggac gcctccgggc cggccatgac cgagatcggc gagcagccgt
     3121 gggggcggga gttcgccctg cgcgacccgg ccggcaactg cgtgcacttc gtggccgagg
     3181 agcaggactg accgacgccg accaacac
//
