LOCUS       sequence-324468-        4710 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway entry plasmid with 3xFlag-pyrin (M694V/FMF variant).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4710)
  AUTHORS   Chirita D, Jamilloux Y, Henry T, Magnotti F
  TITLE     Functional Assessment of Disease-Associated Pyrin Variants.
  JOURNAL   Methods Mol Biol. 2022;2523:179-195. doi: 
            10.1007/978-1-0716-2449-4_12.
  PUBMED    35759198
REFERENCE   2  (bases 1 to 4710)
  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: "2022"; volume: "2523:179-195. doi"; pages: " 
            10.1007/978-1-0716-2449-4_12"
COMMENT     Sequence Label: pENTR1A-3xFlag-M694V-MEFV
FEATURES             Location/Qualifiers
     source          1..4710
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      103..189
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      281..308
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     316..339
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     protein_bind    358..457
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             500..565
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    complement(2939..3038)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3150..3169)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3161..3970
                     /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
                     FQVLEEYPDSGENIVDALAVSLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3228..3247)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4060..4648
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4549..4568
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgctagca tggatctcgg
       61 ggacgtctaa ctactaagcg agagtaggga actgccaggc atcaaataaa acgaaaggct
      121 cagtcggaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt
      181 aggacaaatc cgccgggagc ggatttgaac gttgtgaagc aacggcccgg agggtggcgg
      241 gcaggacgcc cgccataaac tgccaggcat caaactaagc agaaggccat cctgacggat
      301 ggcctttttg cgtttctaca aactcttcct gttagttagt tacttaagct cgggccccaa
      361 ataatgattt tattttgact gatagtgacc tgttcgttgc aacaaattga taagcaatgc
      421 ttttttataa tgccaacttt gtacaaaaaa gcaggcttta aaggaaccaa ttcagtcgac
      481 tggatccggt accaccatgg actacaaaga ccatgacggt gattataaag atcatgacat
      541 cgattacaag gatgacgatg acaagcttgc ggccgcggct aagaccccta gtgaccatct
      601 gctgtccacc ctggaggagc tggtgcccta tgacttcgag aagttcaagt tcaagctgca
      661 gaacaccagt gtgcagaagg agcactccag gatcccccgg agccagatcc agagagccag
      721 gccggtgaag atggccactc tgctggtcac ctactatggg gaagagtacg ccgtgcagct
      781 caccctgcag gtcctgcggg ccatcaacca gcgcctgctg gccgaggagc tccacagggc
      841 agccattcag gaatattcca cacaagaaaa cggcacagat gattccgcag cgtccagctc
      901 cctgggggag aacaagccca ggagcctgaa gactccagac caccccgagg ggaacgaggg
      961 gaacggccct cggccgtacg ggggcggagc tgccagcctg cggtgcagcc agcccgaggc
     1021 cgggaggggg ctgtcgagga agcccctgag caaacgcaga gagaaggcct cggagggcct
     1081 ggacgcgcag ggcaagcctc ggacccggag cccggccctg ccgggcggga gaagccccgg
     1141 cccctgcagg gcgctagagg ggggccaggc cgaggtccgg ctgcgcagaa acgccagctc
     1201 cgcggggagg ctgcaggggc tggcgggggg cgccccgggg cagaaggagt gcaggccctt
     1261 cgaagtgtac ctgccctcgg gaaagatgcg acctagaagc cttgaggtca ccatttctac
     1321 aggggagaag gcgcccgcaa atccagaaat tctcctgact ctagaggaaa agacagctgc
     1381 gaatctggac tcggcaacag aaccccgggc aaggcccact ccggatggag gggcatctgc
     1441 ggacctgaag gaaggccctg gaaatccaga acattcggtc accggaaggc caccagacac
     1501 ggctgcgagt ccccgctgcc acgcccagga aggagaccca gttgacggta cctgtgtgcg
     1561 tgattcctgc agcttccccg aggcagtttc tgggcacccc caggcctcag gcagccgctc
     1621 acctggctgc ccccggtgcc aggactccca tgaaaggaag agcccgggaa gcctaagccc
     1681 ccagcccctg ccacagtgta agcgccacct gaagcaggtc cagctgctct tctgtgagga
     1741 tcacgatgag cccatctgcc tcatctgcag tctgagtcag gagcaccaag gccaccgggt
     1801 gcgccccatt gaggaggtcg ccctggaaca caagaagaaa attcagaagc agctggagca
     1861 tctgaagaag ctgagaaaat caggggagga gcagcgatcc tatggggagg agaaggcagt
     1921 gagctttctg aaacaaactg aagcgctgaa gcagcgggtg cagaggaagc tggagcaggt
     1981 gtactacttc ctggagcagc aagagcattt ctttgtggcc tcactggagg acgtgggcca
     2041 gatggttggg cagatcagga aggcatatga cacccgcgta tcccaggaca tcgccctgct
     2101 cgatgcgctg attggggaac tggaggccaa ggagtgccag tcagaatggg aacttctgca
     2161 ggacattgga gacatcttgc acagggctaa gacagtgcct gtccctgaaa agtggaccac
     2221 tcctcaagag ataaaacaaa agatccaact cctccaccag aagtcagagt ttgtggagaa
     2281 gagcacaaag tacttctcag aaaccctgcg ttcagaaatg gaaatgttca atgttccaga
     2341 gctgattggc gctcaggcac atgctgttaa tgtgattctg gatgcagaaa ccgcttaccc
     2401 caacctcatc ttctctgatg atctgaagag tgttagactt ggaaacaagt gggagaggct
     2461 gcctgatggc ccgcaaagat ttgacagctg tatcattgtt ctgggctctc cgagtttcct
     2521 ctctggccgc cgttactggg aggtggaggt tggagacaag acagcatgga tcctgggagc
     2581 ctgcaagaca tccataagca ggaaagggaa catgactctg tcgccagaga atggctactg
     2641 ggtggtgata atggtgaagg aaaatgagta ccaggcgtcc agcgttcccc cgacccgcct
     2701 gctaataaag gagcctccca agcgtgtggg catcttcgtg gactacagag ttggaagcat
     2761 ctccttttac aatgtgacag ccagatccca catctataca ttcgccagct gctctttctc
     2821 tgggcccctt caacctatct tcagccctgg gacacgtgat ggagggaaga acacagctcc
     2881 tctgactatc tgtccagtgg gtggtcaggg gcctgactga ctgactcgag atatctagac
     2941 ccagctttct tgtacaaagt tggcattata agaaagcatt gcttatcaat ttgttgcaac
     3001 gaacaggtca ctatcagtca aaataaaatc attatttgcc atccagctgc agctctggcc
     3061 cgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat catgaacaat
     3121 aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata ttcaacggga
     3181 aacgtcgagg ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc
     3241 tcgcgataat gtcgggcaat caggtgcgac aatctatcgc ttgtatggga agcccgatgc
     3301 gccagagttg tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat
     3361 ggtcagacta aactggctga cggaatttat gcctcttccg accatcaagc attttatccg
     3421 tactcctgat gatgcatggt tactcaccac tgcgatcccc ggaaaaacag cattccaggt
     3481 attagaagaa tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgtccctgcg
     3541 ccggttgcat tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct
     3601 cgctcaggcg caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga
     3661 gcgtaatggc tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc
     3721 accggattca gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg
     3781 gaaattaata ggttgtattg atgttggacg agtcggaatc gcagaccgat accaggatct
     3841 tgccatccta tggaactgcc tcggtgagtt ttctccttca ttacagaaac ggctttttca
     3901 aaaatatggt attgataatc ctgatatgaa taaattgcag tttcatttga tgctcgatga
     3961 gtttttctaa tcagaattgg ttaattggtt gtaacattat tcagattggg ccccgttcca
     4021 ctgagcgtca gacccggtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     4081 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     4141 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     4201 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     4261 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     4321 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     4381 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     4441 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     4501 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     4561 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     4621 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     4681 ggccttttgc tggccttttg ctcacatgtt
//
