LOCUS       sequence-330901-        3437 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway attB4/attB5 entry clone for mammalian expression using the 
            human EF1alpha promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3437)
  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 3437)
  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-E14: attB4-EF1p>-attB5
FEATURES             Location/Qualifiers
     source          1..3437
                     /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"
     promoter        complement(105..1283)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          114..1052
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(137..157)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1291..1386)
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      1434..1520
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1612..1639
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     rep_origin      complement(1774..2362)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1854..1873)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2505..3314)
                     /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     3228..3247
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3306..3325
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 agctccaaat aatgatttta ttttgactga tagtgacctg ttcgttgcaa caaattgata
       61 agcaatgctt ttttataatg ccaactttgt atacaaaagt tggctcacga cacctgaaat
      121 ggaagaaaaa aactttgaac cactgtctga ggcttgagaa tgaaccaaga tccaaactca
      181 aaaagggcaa attccaagga gaattacatc aagtgccaag ctggcctaac ttcagtctcc
      241 acccactcag tgtggggaaa ctccatcgca taaaacccct ccccccaacc taaagacgac
      301 gtactccaaa agctcgagaa ctaatcgagg tgcctggacg gcgcccggta ctccgtggag
      361 tcacatgaag cgacggctga ggacggaaag gcccttttcc tttgtgtggg tgactcaccc
      421 gcccgctctc ccgagcgccg cgtcctccat tttgagctcc ctgcagcagg gccgggaagc
      481 ggccatcttt ccgctcacgc aactggtgcc gaccgggcca gccttgccgc ccagggcggg
      541 gcgatacacg gcggcgcgag gccaggcacc agagcaggcc ggccagcttg agactacccc
      601 cgtccgattc tcggtggccg cgctcgcagg ccccgcctcg ccgaacatgt gcgctgggac
      661 gcacgggccc cgtcgccgcc cgcggcccca aaaaccgaaa taccagtgtg cagatcttgg
      721 cccgcattta caagactatc ttgccagaaa aaaagcgtcg cagcaggtca tcaaaaattt
      781 taaatggcta gagacttatc gaaagcagcg agacaggcgc gaaggtgcca ccagattcgc
      841 acgcggcggc cccagcgccc aggccaggcc tcaactcaag cacgaggcga aggggctcct
      901 taagcgcaag gcctcgaact ctcccaccca cttccaaccc gaagctcggg atcaagaatc
      961 acgtactgca gccaggtgga agtaattcaa ggcacgcaag ggccataacc cgtaaagagg
     1021 ccaggcccgc gggaaccaca cacggcactt acctgtgttc tggcggcaaa cccgttgcga
     1081 aaaagaacgt tcacggcgac tactgcactt atatacggtt ctcccccacc ctcgggaaaa
     1141 aggcggagcc agtacacgac atcactttcc cagtttaccc cgcgccacct tctctaggca
     1201 ccggttcaat tgccgacccc tccccccaac ttctcgggga ctgtgggcga tgtgcgctct
     1261 gcccactgac gggcaccgga gcctcacggc caacttttct atacaaagtt ggcattataa
     1321 aaaagcattg cttatcaatt tgttgcaacg aacaggtcac tatcagtcaa aataaaatca
     1381 ttatttggag ctccatggta gcgttaacgc gagagtaggg aactgccagg catcaaataa
     1441 aacgaaaggc tcagtcggaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
     1501 ctctcctgag taggacaaat ccgccgggag cggatttgaa cgttgtgaag caacggcccg
     1561 gagggtggcg ggcaggacgc ccgccataaa ctgccaggca tcaaactaag cagaaggcca
     1621 tcctgacgga tggccttttt gcgtttctac aaactcttcc tggctagcgg taatacggtt
     1681 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     1741 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     1801 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     1861 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     1921 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     1981 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2041 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     2101 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     2161 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     2221 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     2281 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     2341 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     2401 gtggaacgaa aactcacgtt aagggatttt ggtcatgagc ttgcgccgtc ccgtcaagtc
     2461 agcgtaatgc tctgccggtg ttacaaccaa ttaaccaatt ctgattagaa aaactcatcg
     2521 agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa
     2581 agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc
     2641 tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg
     2701 tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggcgagaat
     2761 ggcaaaagtt tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca
     2821 tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agcgagacga
     2881 aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg
     2941 aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg
     3001 aatgctgttt ttccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata
     3061 aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca
     3121 tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg
     3181 ggcttcccat acaagcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat
     3241 ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctcga cgtttcccgt
     3301 tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt
     3361 catgatgata tatttttatc ttgtgcaatg taacatcaga gattttgaga cacgggccct
     3421 gcgccctgca gctctag
//
