LOCUS       sequence-146569-        5709 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N-terminal Flag-tagged amino acids 1-90 of NLRP3 (the 
            pyrin domain) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5709)
  AUTHORS   Shi H, Wang Y, Li X, Zhan X, Tang M, Fina M, Su L, Pratt D, Bu CH, 
            Hildebrand S, Lyon S, Scott L, Quan J, Sun Q, Russell J, Arnett S, 
            Jurek P, Chen D, Kravchenko VV, Mathison JC, Moresco EM, Monson NL, 
            Ulevitch RJ, Beutler B
  TITLE     NLRP3 activation and mitosis are mutually exclusive events 
            coordinated by NEK7, a new inflammasome component.
  JOURNAL   Nat Immunol. 2016 Mar;17(3):250-8. doi: 10.1038/ni.3333. Epub 2015 
            Dec 7.
  PUBMED    26642356
REFERENCE   2  (bases 1 to 5709)
  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: "10.1038/ni.3333"; 
            journalName: "Nat Immunol"; date: "2016-03"; volume: "17"; issue: 
            "3"; pages: "250-8"
COMMENT     Sequence Label: pcDNA3-N-Flag-NLRP3 1-90
FEATURES             Location/Qualifiers
     source          1..5709
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             922..945
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(1261..1279)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1262..1279)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1299..1316)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1305..1529
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1575..2003
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1662..1681)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1872..1893
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2012..2032)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2017..2347
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2198..2333
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2260..2279
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2414..3208
                     /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(2468..2487)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3078..3097
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3382..3503
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3419..3438)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3473..3492
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3552..3568)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3552..3568)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3565..3587)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3576..3592
                     /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)."
     promoter        complement(join(3600..3606,3607..3624,3625..3630))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3645..3666
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3783..3800)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3954..4542)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4034..4053)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4713..5504,5505..5573))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5336..5355
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5574..5678)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat cgcccaccat ggactacaaa gacgatgacg ataaaatcga tatcgaattc
      961 atgacgagtg tccgttgcaa gctggctcag tatctagagg accttgaaga tgtggacctc
     1021 aagaaattca aaatgcattt ggaagattac ccgcccgaga aaggctgtat cccagtcccc
     1081 aggggccaga tggagaaggc agatcacttg gatctagcca cactcatgat tgacttcaat
     1141 ggtgaggaga aggcctgggc catggctgtg tggatctttg ctgcgatcaa caggcgagac
     1201 ctctgggaaa aagctaagaa ggaccagcca tagctcgagc atgcatctag agggccctat
     1261 tctatagtgt cacctaaatg ctagagctcg ctgatcagcc tcgactgtgc cttctagttg
     1321 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     1381 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     1441 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     1501 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     1561 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     1621 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     1681 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg gcatcccttt
     1741 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     1801 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     1861 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     1921 ttcttttgat ttataaggga ttttggggat ttcggcctat tggttaaaaa atgagctgat
     1981 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
     2041 tccccaggct ccccaggcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     2101 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     2161 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     2221 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     2281 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     2341 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg
     2401 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     2461 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     2521 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     2581 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     2641 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     2701 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     2761 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     2821 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     2881 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     2941 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     3001 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     3061 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     3121 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     3181 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     3241 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     3301 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     3361 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     3421 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3481 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     3541 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3601 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3661 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3721 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     3781 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3841 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3901 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3961 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4021 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4081 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4141 ctttctcaat gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4201 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4261 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4321 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4381 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4441 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4501 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4561 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4621 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4681 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4741 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4801 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4861 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4921 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4981 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     5041 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5101 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5161 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5221 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5281 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5341 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5401 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5461 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5521 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5581 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5641 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     5701 cctgacgtc
//
