LOCUS       sequence-333338-        3150 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway attB4/attB1r entry clone for baculovirus expression of dual 
            Strep2 tagged proteins using the polyhedrin promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3150)
  AUTHORS   Wall VE, Garvey LA, Mehalko JL, Procter LV, Esposito D
  TITLE     Combinatorial assembly of clone libraries using site-specific 
            recombination.
  JOURNAL   Methods Mol Biol. 2014;1116:193-208. doi: 
            10.1007/978-1-62703-764-8_14.
  PUBMED    24395366
REFERENCE   2  (bases 1 to 3150)
  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: "Methods Mol
            Biol."; date: "2014"; volume: "1116:193-208. doi"; pages: " 
            10.1007/978-1-62703-764-8_14"
COMMENT     Sequence Label: C417-E12: attB4-polyhedrinp>Strep2x2-nostop-at...
FEATURES             Location/Qualifiers
     source          1..3150
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(34..120)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     169..191
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     183..200
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     184..200
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    256..351
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        417..508
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     433..450
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             join(515..538,539..574,575..598)
                     /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"
     protein_bind    602..726
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(798..817)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(799..817)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(822..838)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(822..838)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1263..2054
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      2147..2735
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2636..2655
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2889..2906
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3065..3092)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttgcttcaca acgttcaaat ccgctcccgg cggatttgtc ctactcagga gagcgttcac
       61 cgacaaacaa cagataaaac gaaaggccca gtcttccgac tgagcctttc gttttatttg
      121 atgcctggca gttccctact ctcgcgttaa cgctagcatg gatgttttcc cagtcacgac
      181 gttgtaaaac gacggccagt cttaaaattc gagctcggtc cgatcgggcc cgcgttaacg
      241 ctaccatgga gctccaaata atgattttat tttgactgat agtgacctgt tcgttgcaac
      301 aaattgataa gcaatgcttt tttataatgc caactttgta tagaaaagtt ggcgttcgcc
      361 caggactcta gctatagttc tagtggttgg ctacgtatac tccggaatat taatagatca
      421 tggagataat taaaatgata accatctcgc aaataaataa gtattttact gttttcgtaa
      481 cagttttgta ataaaaaaac ctataaatat gggctggagt caccctcaat ttgagaaggg
      541 agggggttct ggtggcggat ccggcggatc agcctggagc catccacagt tcgaaaaggg
      601 aacaactttg tacaaaaaag ttgaacgaga aacgtaaaat gatataaata tcaatatatt
      661 aaattagatt ttgcataaaa aacagactac ataatactgt aaaacacaac atatgcagtc
      721 actatgaacc aactacttag atggtattag tgacctgtag aattcgagct ctagagctgc
      781 agggcggccg cgatatcccc tatagtgagt cgtattacat ggtcatagct gtttcctggc
      841 agctctggcc cgtgtctcaa aatctctgat gttacattgc acaagatcgg tacccgggga
      901 tcctctagac cagccaggac agaaatgcct cgacttcgct gctacccaag gttgccgggt
      961 gacgcacacc gtggaaacgg atgaaggcac gaacccagtg gacataagcc tgttcggttc
     1021 gtaagctgta atgcaagtag cgtatgcgct cacgcaactg gtccagaacc ttgaccgaac
     1081 gcagcggtgg taacggcgca gtggcggttt tcatggcttg ttatgactgt ttttttgggg
     1141 tacagtctat gcctcgggca tccaagcagc aagcgcgtta cgccgtgggt cgatgtttga
     1201 tgttatggag cagcaacgat gttacgcagc agggcagtcg ccctaaaaca aagttaaaca
     1261 ttatgaggga agcggtgatc gccgaagtat cgactcaact atcagaggta gttggcgtca
     1321 tcgagcgcca tctcgaaccg acgttgctgg ccgtacattt gtacggctcc gcagtggatg
     1381 gcggcctgaa gccacacagt gatattgatt tgctggttac ggtgaccgta aggcttgatg
     1441 aaacaacgcg gcgagctttg atcaacgacc ttttggaaac ttcggcttcc cctggagaga
     1501 gcgagattct ccgcgctgta gaagtcacca ttgttgtgca cgacgacatc attccgtggc
     1561 gttatccagc taagcgcgaa ctgcaatttg gagaatggca gcgcaatgac attcttgcag
     1621 gtatcttcga gccagccacg atcgacattg atctggctat cttgctgaca aaagcaagag
     1681 aacatagcgt tgccttggta ggtccagcgg cggaggaact ctttgatccg gttcctgaac
     1741 aggatctatt tgaggcgcta aatgaaacct taacgctatg gaactcgccg cccgactggg
     1801 ctggcgatga gcgaaatgta gtgcttacgt tgtcccgcat ttggtacagc gcagtaaccg
     1861 gcaaaatcgc gccgaaggat gtcgctgccg actgggcaat ggagcgcctg ccggcccagt
     1921 atcagcccgt catacttgaa gctagacagg cttatcttgg acaagaagaa gatcgcttgg
     1981 cctcgcgcgc agatcagttg gaagaatttg tccactacgt gaaaggcgag atcaccaagg
     2041 tagtcggcaa ataaccctcg agccacccat gaccaaaatc ccttaacgtg agttacgcgt
     2101 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     2161 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     2221 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     2281 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     2341 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     2401 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     2461 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     2521 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     2581 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     2641 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     2701 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     2761 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     2821 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     2881 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     2941 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
     3001 cgggcagtga gcgcaacgca attaatacgc gtaccgctag ccaggaagag tttgtagaaa
     3061 cgcaaaaagg ccatccgtca ggatggcctt ctgcttagtt tgatgcctgg cagtttatgg
     3121 cgggcgtcct gcccgccacc ctccgggccg
//
