LOCUS       sequence-175061-        5689 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP with Frt sites for FLP-mediated knock-out of gene function in C.
            elegans.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5689)
  AUTHORS   Munoz-Jimenez C, Ayuso C, Dobrzynska A, Torres-Mendez A, Ruiz PC, 
            Askjaer P
  TITLE     An Efficient FLP-Based Toolkit for Spatiotemporal Control of Gene 
            Expression in Caenorhabditis elegans.
  JOURNAL   Genetics. 2017 Aug;206(4):1763-1778. doi: 
            10.1534/genetics.117.201012. Epub 2017 Jun 23.
  PUBMED    28646043
REFERENCE   2  (bases 1 to 5689)
  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"; doi: 
            "10.1534/genetics.117.201012"; journalName: "Genetics"; date: 
            "2017-08"; volume: "206"; issue: "4"; pages: "1763-1778"
COMMENT     Sequence Label: pBN312
FEATURES             Location/Qualifiers
     source          1..5689
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    6..105
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(144..172)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     protein_bind    311..344
                     /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)"
     protein_bind    550..583
                     /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)"
     protein_bind    799..832
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             2796..2807
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    2956..2989
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     3179..3199
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     protein_bind    complement(3251..3350)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3367..3386)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3368..3386)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3391..3407)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3391..3407)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3509..3528)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3520..4329
                     /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(3587..3606)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4476..5064
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4965..4984
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5218..5235
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(5394..5421)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(5513..5599)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     5648..5670
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5662..5679
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5663..5679
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ggccccaaat aatgatttta ttttgactga tagtgacctg ttcgttgcaa caaattgatg
       61 agcaatgctt ttttataatg ccaactttgt acaaaaaagc aggctgctct tcgatgagca
      121 aaggagaaga acttttcact ggagttgtcc caattcttgt tgaattagat ggtgatgtta
      181 atgggcacaa attttctgtc agtggagagg gtgaaggtga tgcaacatac ggaaaactta
      241 cccttaaatt tatttgcact actggaaaac tacctgttcc atgggtaagt ttaaacatat
      301 atatactaac gaagttccta ttctctagaa agtataggaa cttctaaccc tgattattta
      361 aattttcagc caacacttgt cactactttc tgttatggtg ttcaatgctt ctcgagatac
      421 ccagatcata tgaaacggca tgactttttc aagagtgcca tgcccgaagg ttatgtacag
      481 gaaagaacta tatttttcaa agatgacggg aactacaaga cacgtaagtt taaacagttc
      541 ggtactaacg aagttcctat tctctagaaa gtataggaac ttctaaccat acatatttaa
      601 attttcaggt gctgaagtca agtttgaagg tgataccctt gttaatagaa tcgagttaaa
      661 aggtattgat tttaaagaag atggaaacat tcttggacac aaattggaat acaactataa
      721 ctcacacaat gtatacatca tggcagacaa acaaaagaat ggaatcaaag ttgtaagttt
      781 aaacatgatt ttactaacat aacttcgtat aatgtatgct atacgaagtt attactagac
      841 attctctaat gaaaaaatct ttcagttgaa attgaaaatg agttaaagtt ggagttttta
      901 ttgaaaacag atttccgtgt gattagtgtt tttagcgagt gtgacaggac agcgaaaaaa
      961 tatagaaaca aggggggaac tgaaaagctt aggaatgcat tgaacatgag aaggggaaag
     1021 ggaaggaaca aactagacag gaattattgg aatttaatca catttggagt tttttttcta
     1081 ttcgacagaa taattatcca gaacattttt gtattaaata tttatgcatc atatgagtag
     1141 tcggctttgt tgtgcatgac gagtttgtta tcgacgaaat agaagctgtc agaacgagtc
     1201 tcgtttggat tgttgatcat gtcgtccact gaaaaagaga ttagtctttt gaattgtact
     1261 ttttagaata atgactcact gagttgttga gagagttgag ggaactcata gatatgttca
     1321 cagttgtttc gtgaattcgg aatacagaat ccgaattcaa agtcaaaaca cttcagaagg
     1381 cgatctttga agaagtgacg ttcgatcatt cggaaatgat ggattgggat gtctcctact
     1441 ttgaattcca cagttgctcc gaccgttttg agtttcaaaa agtttggagc aaatctataa
     1501 cgcacgtatc ttgccgactc ctgtggcgac tcatcattct cctgatcgtt ctccggtttg
     1561 gcgatctcga aaagcacttg ctcagtgtcc agatcacgga tttggaactt ggtgaactcg
     1621 atgttataga tgttcgcaga tggggagcat aagaatccta aatttatgtt ttaaactgaa
     1681 atccaaaggg agcaagatac cttgagtgat tcccggaagt gctaaaacgt cgttcggagt
     1741 gatttgagct ttcttcgcaa gctccgattc cgttgtgatt ccttgttcgg tgcttggtgg
     1801 tggccgtggc atctggaaat atggaaaagt tcaacaaaaa gaaaagagaa aagaatgaaa
     1861 tcggatatca agagttagtt gagcggtttc tctagttttc tgagtctcac ctgcgacggg
     1921 aaggtcgccg agccgggtgg aatcgatcgt tgttgctcgg ctttcatatc ggtttggttg
     1981 gaagcggctg aaaacggaaa gaagtggaag aaggaaaaga gtgtggtgtg acaggaaaat
     2041 ggtaattaga gggtgccaaa taaccagcta tattttgttt ttttttgaaa acatttttaa
     2101 aaagaaaaat acgataatga tatcagatgg atttccggaa aactggtatg aaaaatttca
     2161 acctttttga gtacatgtaa tcaaaataca ctttgtaaat tatcattttt attgaaactc
     2221 caccattttt ctatttataa cgctaataat ttgaaaaaga aacctattgc gaaccgcggg
     2281 gtgaatccca aaaacgaatg cgttttggtg gagtgattga ttcgaatcga agaagaaaaa
     2341 gaagaagacg tggaatagag agctcactct taaccgagca gcacacaccg acagaaaaaa
     2401 aaatgaaatg aatgagggtc ttcttcttct tcttcttcga atgattgaca gaaatgggaa
     2461 aaagaggaag attgagaagg gaaaaaggaa ggagaaaaga agcagaagaa gacgtcagag
     2521 aggagaggaa cgagcggaaa agcagcgggc gcaagtcata gaagtagcag agctggggag
     2581 aagaagacac tatccaagaa aggaatgacg agagagtatg caaaggggta tagggtgcag
     2641 acagaatagg aacagaataa cagatgatga gccaagaaga gttgaaaagg gcgatgaatt
     2701 tgtcatgtaa cttaatttgg gtcaatttga gcatgatgaa ttgaaatcat cccttgttgg
     2761 gagttaataa ccggtttgtt atcagaaacc ctgtaataga agggcgccct aactttgagc
     2821 caattcatcc cggtttctgt caaatatatc aaaaagtggt caactgacaa attgtttttg
     2881 atattataat aaacatttta tccgttaaca attttcgaat actttttaca aggacttgga
     2941 taaattggct caaagataac ttcgtataat gtatgctata cgaagttatt aactaatctg
     3001 atttaaattt tcagaacttc aaaattagac acaacattga agatggaagc gttcaactag
     3061 cagaccatta tcaacaaaat actccaattg gcgatggccc tgtcctttta ccagacaacc
     3121 attacctgtc cacacaatct gccctttcga aagatcccaa cgaaaagaga gaccacatgg
     3181 tccttcttga gtttgtaaca gctgctggga ttacacatgg catggatgaa ctatacaaag
     3241 gtcgaagagc acccagcttt cttgtacaaa gttggcatta taagaaagca ttgcttatca
     3301 atttgttgca acgaacaggt cactatcagt caaaataaaa tcattatttg ccatccagct
     3361 gatatcccct atagtgagtc gtattacatg gtcatagctg tttcctggca gctctggccc
     3421 gtgtctcaaa atctctgatg ttacattgca caagataaaa atatatcatc atgaacaata
     3481 aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa
     3541 acgtcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct
     3601 cgcgataatg tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg
     3661 ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg
     3721 gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt
     3781 actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc attccaggta
     3841 ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc
     3901 cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc
     3961 gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt tgatgacgag
     4021 cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt gccattctca
     4081 ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg
     4141 aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt
     4201 gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcaa
     4261 aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat gctcgatgag
     4321 tttttctaat cagaattggt taattggttg taacactggc agagcattac gctgacttga
     4381 cgggacggcg caagctcatg accaaaatcc cttaacgtga gttacgcgtc gttccactga
     4441 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     4501 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     4561 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     4621 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     4681 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     4741 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     4801 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     4861 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     4921 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     4981 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     5041 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     5101 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     5161 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     5221 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
     5281 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     5341 cgcaacgcaa ttaatacgcg taccgctagc caggaagagt ttgtagaaac gcaaaaaggc
     5401 catccgtcag gatggccttc tgcttagttt gatgcctggc agtttatggc gggcgtcctg
     5461 cccgccaccc tccgggccgt tgcttcacaa cgttcaaatc cgctcccggc ggatttgtcc
     5521 tactcaggag agcgttcacc gacaaacaac agataaaacg aaaggcccag tcttccgact
     5581 gagcctttcg ttttatttga tgcctggcag ttccctactc tcgcgttaac gctagcatgg
     5641 atgttttccc agtcacgacg ttgtaaaacg acggccagtc ttaagctcg
//
