LOCUS       sequence-330601-        3038 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway attB5r/attB1r entry clone for generation of N-terminal 
            3xFLAG-tagged fusion proteins.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3038)
  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 3038)
  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: C514-E23: attB5r-RU5u-3xFLAG-nostop-attB1r
FEATURES             Location/Qualifiers
     source          1..3038
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(14..100)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    complement(185..308)
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     LTR             312..580
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             593..658
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    665..789
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(861..880)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(862..880)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(885..901)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(885..901)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1326..2117
                     /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      2210..2798
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2699..2718
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2933..2960)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ccgctcccgg cggatttgtc ctactcagga gagcgttcac cgacaaacaa cagataaaac
       61 gaaaggccca gtcttccgac tgagcctttc gttttatttg atgcctggca gttccctact
      121 ctcgcgttaa cgctaccatg gagctcgaat tctacagtca ctaataccat ctaagtagtt
      181 gattcatagt gactgcatat gttgtgtttt acagtattat gtagtctgtt ttttatgcaa
      241 aatctaattt aatatattga tatttatatc attttacgtt tctcgttcaa cttttgtata
      301 caaagtggcg aggggctcgc atctctcctt cacgcgcccg ccgccctacc tgaggccgcc
      361 atccacgccg gttgagtcgc gttctgccgc ctcccgcctg tggtgcctcc tgaactgcgt
      421 ccgccgtcta ggtaagttta aagctcaggt cgagaccggg cctttgtccg gcgctccctt
      481 ggagcctacc tagactcagc cggctctcca cgctttgcct gaccctgctt gctcaactct
      541 acgtctttgt ttcgttttct gttctgcgcc gttacagatc ggcaccatgg gcgactacaa
      601 agaccacgac ggtgattata aggatcatga catcgattat aaggatgacg atgacaaggg
      661 cagcacaact ttgtacaaaa aagttgaacg agaaacgtaa aatgatataa atatcaatat
      721 attaaattag attttgcata aaaaacagac tacataatac tgtaaaacac aacatatgca
      781 gtcactatga accaactact tagatggtat tagtgacctg tagaattcga gctctagagc
      841 tgcagggcgg ccgcgatatc ccctatagtg agtcgtatta catggtcata gctgtttcct
      901 ggcagctctg gcccgtgtct caaaatctct gatgttacat tgcacaagat cggtacccgg
      961 ggatcctcta gaccagccag gacagaaatg cctcgacttc gctgctaccc aaggttgccg
     1021 ggtgacgcac accgtggaaa cggatgaagg cacgaaccca gtggacataa gcctgttcgg
     1081 ttcgtaagct gtaatgcaag tagcgtatgc gctcacgcaa ctggtccaga accttgaccg
     1141 aacgcagcgg tggtaacggc gcagtggcgg ttttcatggc ttgttatgac tgtttttttg
     1201 gggtacagtc tatgcctcgg gcatccaagc agcaagcgcg ttacgccgtg ggtcgatgtt
     1261 tgatgttatg gagcagcaac gatgttacgc agcagggcag tcgccctaaa acaaagttaa
     1321 acattatgag ggaagcggtg atcgccgaag tatcgactca actatcagag gtagttggcg
     1381 tcatcgagcg ccatctcgaa ccgacgttgc tggccgtaca tttgtacggc tccgcagtgg
     1441 atggcggcct gaagccacac agtgatattg atttgctggt tacggtgacc gtaaggcttg
     1501 atgaaacaac gcggcgagct ttgatcaacg accttttgga aacttcggct tcccctggag
     1561 agagcgagat tctccgcgct gtagaagtca ccattgttgt gcacgacgac atcattccgt
     1621 ggcgttatcc agctaagcgc gaactgcaat ttggagaatg gcagcgcaat gacattcttg
     1681 caggtatctt cgagccagcc acgatcgaca ttgatctggc tatcttgctg acaaaagcaa
     1741 gagaacatag cgttgccttg gtaggtccag cggcggagga actctttgat ccggttcctg
     1801 aacaggatct atttgaggcg ctaaatgaaa ccttaacgct atggaactcg ccgcccgact
     1861 gggctggcga tgagcgaaat gtagtgctta cgttgtcccg catttggtac agcgcagtaa
     1921 ccggcaaaat cgcgccgaag gatgtcgctg ccgactgggc aatggagcgc ctgccggccc
     1981 agtatcagcc cgtcatactt gaagctagac aggcttatct tggacaagaa gaagatcgct
     2041 tggcctcgcg cgcagatcag ttggaagaat ttgtccacta cgtgaaaggc gagatcacca
     2101 aggtagtcgg caaataaccc tcgagccacc catgaccaaa atcccttaac gtgagttacg
     2161 cgtcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     2221 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     2281 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     2341 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     2401 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     2461 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     2521 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     2581 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     2641 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     2701 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     2761 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     2821 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     2881 attctgtgga taaccgtatt accgctagcc aggaagagtt tgtagaaacg caaaaaggcc
     2941 atccgtcagg atggccttct gcttagtttg atgcctggca gtttatggcg ggcgtcctgc
     3001 ccgccaccct ccgggccgtt gcttcacaac gttcaaat
//
