LOCUS       sequence-330612-        2440 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway attB4/attB1r entry clone for bacterial expression using the 
            bacteriophage T7 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2440)
  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 2440)
  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: C453-E26: attB4-T7p>-attB5
FEATURES             Location/Qualifiers
     source          1..2440
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    7..102
                     /gene="mutant version of attL"
                     /label=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    118..142
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     complement(142..161)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(143..161)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(249..268)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     protein_bind    complement(294..389)
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      437..523
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      615..642
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     rep_origin      complement(777..1365)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(857..876)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1508..2317)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2231..2250
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2309..2328
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 agctccaaat aatgatttta ttttgactga tagtgacctg ttcgttgcaa caaattgata
       61 agcaatgctt ttttataatg ccaactttgt atacaaaagt tggcttattt ctagagggga
      121 attgttatcc gctcacaatt cccctatagt gagtcgtatt aatttcgcgg gatcgagatc
      181 gatctcgatc ctctacgccg gacgcatcgt ggccggcatc accggcgcca caggtgcggt
      241 tgctggcgcc tatatcgccg acatcaccga tggggaagat cgggctcgcc agccaacttt
      301 tctatacaaa gttggcatta taaaaaagca ttgcttatca atttgttgca acgaacaggt
      361 cactatcagt caaaataaaa tcattatttg gagctccatg gtagcgttaa cgcgagagta
      421 gggaactgcc aggcatcaaa taaaacgaaa ggctcagtcg gaagactggg cctttcgttt
      481 tatctgttgt ttgtcggtga acgctctcct gagtaggaca aatccgccgg gagcggattt
      541 gaacgttgtg aagcaacggc ccggagggtg gcgggcagga cgcccgccat aaactgccag
      601 gcatcaaact aagcagaagg ccatcctgac ggatggcctt tttgcgtttc tacaaactct
      661 tcctggctag cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
      721 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
      781 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
      841 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
      901 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
      961 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     1021 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     1081 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     1141 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     1201 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     1261 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     1321 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     1381 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     1441 agcttgcgcc gtcccgtcaa gtcagcgtaa tgctctgccg gtgttacaac caattaacca
     1501 attctgatta gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat
     1561 tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc
     1621 agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa
     1681 tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag
     1741 tgacgactga atccggcgag aatggcaaaa gtttatgcat ttctttccag acttgttcaa
     1801 caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc
     1861 gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag
     1921 gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat
     1981 caggatattc ttctaatacc tggaatgctg tttttccggg gatcgcagtg gtgagtaacc
     2041 atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca
     2101 gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt
     2161 tcagaaacaa ctctggcgca tcgggcttcc catacaagcg atagattgtc gcacctgatt
     2221 gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta
     2281 atcgcggcct cgacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta
     2341 tgtaagcaga cagttttatt gttcatgatg atatattttt atcttgtgca atgtaacatc
     2401 agagattttg agacacgggc cctgcgccct gcagctctag
//
