LOCUS       sequence-147260-        7296 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N-terminal Flag-tagged amino acids 91-710 of NLRP3 (the 
            nucleotide-binding domain) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7296)
  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 7296)
  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 91-710
FEATURES             Location/Qualifiers
     source          1..7296
                     /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(2848..2866)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2849..2866)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2886..2903)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2892..3116
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3162..3590
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3249..3268)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3459..3480
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3599..3619)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3604..3934
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3785..3920
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3847..3866
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4001..4795
                     /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(4055..4074)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4665..4684
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4969..5090
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5006..5025)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5060..5079
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5139..5155)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5139..5155)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5152..5174)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5163..5179
                     /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(5187..5193,5194..5211,5212..5217))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5232..5253
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5370..5387)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5541..6129)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5621..5640)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6300..7091,7092..7160))
                     /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     6923..6942
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7161..7265)
                     /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 gagtggaatg acacgtgtac atcacattcc tctatggtat gccaggagga cagccttgaa
     1021 gaagagtgga tgggtttgct gggatatctc tcccgcatct ccatttgtaa aaagaagaaa
     1081 gattactgta agatgtacag acgacatgtg agaagcaggt tctactctat caaggacagg
     1141 aacgcgcgtc taggtgagag tgtggacctc aacagtcgct acacgcagct ccaactggtc
     1201 aaggagcatc caagcaagca ggagcgggag catgaactcc tgaccatcgg ccggactaaa
     1261 atgcgggaca gccccatgag ttcccttaag ctggagctgc tgtttgagcc cgaggacggg
     1321 cactcggagc ctgtgcacac agtggtgttc cagggagcag caggcatcgg gaaaaccatc
     1381 ctagccagga agattatgtt ggactgggca ctgggaaagc tcttcaaaga caaatttgac
     1441 tatttgttct ttatccactg ccgagaggtg agcctcagga cgccaaggag tctagcagac
     1501 ctgattgtca gctgctggcc tgacccaaac ccaccagtgt gcaagatcct gcgcaagcct
     1561 tccaggatcc tcttcctcat ggatggcttt gatgagctac aaggggcctt tgacgagcac
     1621 attggggagg tctgcacaga ctggcaaaag gctgtgcggg gagacattct gctaagcagc
     1681 ctcatccgaa agaaactgct gcccaaggcc tctctgctca taacgacgag gccggtagcc
     1741 ttggagaaac tgcagcatct cctggaccac ccccgccatg tggagatcct aggtttctct
     1801 gaggccaaaa ggaaggagta tttctttaag tatttctcca acgagctgca ggcccgggag
     1861 gccttcaggc tgatccaaga gaatgaggtc ctctttacca tgtgcttcat ccccctggtc
     1921 tgctggattg tgtgcacggg gctaaagcaa cagatggaga ccgggaagag cctggcccag
     1981 acctccaaga ccactacggc cgtctacgtc ttcttccttt ccagcctgct gcaatcccgg
     2041 gggggcattg aggagcatct cttctctgac tacctacagg ggctctgttc actggctgcg
     2101 gatggaattt ggaaccagaa aatcctattt gaggagtgtg atctgcggaa gcacggcctg
     2161 cagaagactg acgtctccgc tttcctgagg atgaacgtgt tccagaagga agtggactgc
     2221 gagagattct acagcttcag ccacatgact ttccaggagt tcttcgctgc tatgtactat
     2281 ttgctggaag aggaggcaga gggggagacc gtgaggaaag gaccaggagg ttgttcagat
     2341 cttctgaacc gagacgtgaa ggtcctacta gaaaattacg gcaagtttga aaaaggctat
     2401 ctgatttttg ttgtccgatt cctctttggc cttgtaaacc aggagagaac ctcttatttg
     2461 gagaagaaac taagttgcaa gatctctcag caagtcagac tggaactact gaagtggatt
     2521 gaagtgaaag ccaaggccaa gaagctgcag tggcagccca gccaactgga actgttctac
     2581 tgcctgtacg agatgcagga ggaagacttt gtgcagagtg ccatggacca ctttcccaaa
     2641 attgagatca acctctctac cagaatggac cacgtggttt cctccttttg tattaagaac
     2701 tgtcataggg tcaaaacgct ttccctgggt ttttttcaca actcgcccaa ggaggaagaa
     2761 gaagagagga gaggaggtcg acccttggac caggttcagt gtgttttccc agacactcat
     2821 ctcgagcatg catctagagg gccctattct atagtgtcac ctaaatgcta gagctcgctg
     2881 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     2941 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     3001 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     3061 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     3121 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     3181 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     3241 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     3301 tcaagctcta aatcggggca tccctttagg gttccgattt agtgctttac ggcacctcga
     3361 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     3421 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     3481 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tggggatttc
     3541 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     3601 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc caggcaggca gaagtatgca
     3661 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     3721 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     3781 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     3841 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     3901 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     3961 tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     4021 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     4081 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     4141 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     4201 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     4261 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     4321 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     4381 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     4441 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     4501 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     4561 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     4621 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     4681 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     4741 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     4801 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     4861 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     4921 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     4981 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     5041 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     5101 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     5161 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     5221 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     5281 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5341 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     5401 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     5461 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     5521 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     5581 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     5641 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     5701 gcctttctcc cttcgggaag cgtggcgctt tctcaatgct cacgctgtag gtatctcagt
     5761 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     5821 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     5881 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     5941 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     6001 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     6061 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     6121 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     6181 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     6241 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6301 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6361 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6421 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6481 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6541 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6601 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6661 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     6721 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     6781 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     6841 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     6901 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     6961 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     7021 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     7081 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     7141 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     7201 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     7261 ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
