LOCUS       sequence-165133-        3860 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3' entry vector containing FRT-flanked kanamycin resistance cassette
            (KanR) with pA for FLP-induced kanamycin resistance, drug selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3860)
  AUTHORS   Fowler DK, Stewart S, Seredick S, Eisen JS, Stankunas K, Washbourne 
            P
  TITLE     A MultiSite Gateway Toolkit for Rapid Cloning of Vertebrate 
            Expression Constructs with Diverse Research Applications.
  JOURNAL   PLoS One. 2016 Aug 8;11(8):e0159277. doi: 
            10.1371/journal.pone.0159277. eCollection 2016.
  PUBMED    27500400
REFERENCE   2  (bases 1 to 3860)
  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: "PLoS One.";
            date: "2016-08-8"; volume: "11(8):e0159277. doi"; pages: " 
            10.1371/journal.pone.0159277. eCollection 2016"
COMMENT     Sequence Label: p3E-FRT-KanR-FRT pA
FEATURES             Location/Qualifiers
     source          1..3860
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(624..747)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(755..788)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             complement(789..1583)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(900..919)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1510..1529
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     protein_bind    complement(1737..1770)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     polyA_signal    1772..1906
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1796..1815)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1850..1869
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    complement(1969..2064)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2101..2120)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2102..2120)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2125..2141)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2125..2141)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2243..2262)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2254..3063
                     /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     complement(2321..2340)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3210..3798
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3699..3718
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggccctg cagctctaga gctcgaattc tacaggtcac
      601 taataccatc taagtagttg gttcatagtg actgcatatg ttgtgtttta cagtattatg
      661 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt
      721 ctcgttcaac tttcttgtac aaagtggggg atccgaagtt cctatacttt ctagagaata
      781 ggaacttctc agaagaactc gtcaagaagg cgatagaagg cgatgcgctg cgaatcggga
      841 gcggcgatac cgtaaagcac gaggaagcgg tcagcccatt cgccgccaag ctcttcagca
      901 atatcacggg tagccaacgc tatgtcctga tagcggtccg ccacacccag ccggccacag
      961 tcgatgaatc cagaaaagcg gccattttcc accatgatat tcggcaagca ggcatcgcca
     1021 tgggtcacga cgagatcctc gccgtcgggc atgcgcgcct tgagcctggc gaacagttcg
     1081 gctggcgcga gcccctgatg ctcttcgtcc agatcatcct gatcgacaag accggcttcc
     1141 atccgagtac gtgctcgctc gatgcgatgt ttcgcttggt ggtcgaatgg gcaggtagcc
     1201 ggatcaagcg tatgcagccg ccgcattgca tcagccatga tggatacttt ctcggcagga
     1261 gcaaggtggg atgacaggag atcctgcccc ggcacttcgc ccaatagcag ccagtccctt
     1321 cccgcttcag tgacaacgtc gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac
     1381 gatagccgcg ctgcctcgtc ctgcagttca ttcagggcac cggacaggtc ggtcttgaca
     1441 aaaagaaccg ggcgcccctg cgctgacagc cggaacacgg cggcatcaga gcagccgatt
     1501 gtctgttgtg cccagtcata gccgaatagc ctctccaccc aagcggccgg agaacctgcg
     1561 tgcaatccat cttgttcaat catgcgaaac gatcctcatc ctgtctcttg atcagatctt
     1621 gatcccctgc gccatcagat ccttggcggc aagaaagcca tccagtttac tttgcagggc
     1681 ttcccaacct taccagaggg cgccccagct ggcaattccg gttcgcttgc tgtccagaag
     1741 ttcctatact ttctagagaa taggaacttc ggatccagac atgataagat acattgatga
     1801 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     1861 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     1921 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttcca actttattat
     1981 acaaagttgg cattataaaa aagcattgct tatcaatttg ttgcaacgaa caggtcacta
     2041 tcagtcaaaa taaaatcatt atttggagct ccatggtagc gttaacgcgg ccgcgatatc
     2101 ccctatagtg agtcgtatta catggtcata gctgtttcct ggcagctctg gcccgtgtct
     2161 caaaatctct gatgttacat tgcacaagat aaaaatatat catcatgaac aataaaactg
     2221 tctgcttaca taaacagtaa tacaaggggt gttatgagcc atattcaacg ggaaacgtcg
     2281 aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat
     2341 aatgtcgggc aatcaggtgc gacaatctat cgcttgtatg ggaagcccga tgcgccagag
     2401 ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga
     2461 ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct
     2521 gatgatgcat ggttactcac cactgcgatc cccggaaaaa cagcattcca ggtattagaa
     2581 gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg
     2641 cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg tctcgctcag
     2701 gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga cgagcgtaat
     2761 ggctggcctg ttgaacaagt ctggaaagaa atgcataaac ttttgccatt ctcaccggat
     2821 tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga ggggaaatta
     2881 ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga tcttgccatc
     2941 ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt tcaaaaatat
     3001 ggtattgata atcctgatat gaataaattg cagtttcatt tgatgctcga tgagtttttc
     3061 taatcagaat tggttaattg gttgtaacac tggcagagca ttacgctgac ttgacgggac
     3121 ggcgcaagct catgaccaaa atcccttaac gtgagttacg cgtcgttcca ctgagcgtca
     3181 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     3241 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     3301 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     3361 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     3421 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     3481 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     3541 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     3601 cattgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     3661 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     3721 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     3781 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     3841 tggccttttg ctcacatgtt
//
