LOCUS       sequence-192740-        6017 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N-terminal Flag-tagged ASC1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6017)
  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 6017)
  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-ASC1
FEATURES             Location/Qualifiers
     source          1..6017
                     /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(1570..1588)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1571..1588)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1608..1625)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1614..1838
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1884..2312
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1971..1990)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2181..2202
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2321..2341)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2326..2655
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2506..2641
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2568..2587
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2722..3516
                     /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(2776..2795)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3386..3405
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3690..3811
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3727..3746)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3781..3800
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3860..3876)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3860..3876)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3873..3895)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3884..3900
                     /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(3908..3914,3915..3932,3933..3938))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3953..3974
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4091..4108)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4262..4850)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4342..4361)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5021..5812,5813..5881))
                     /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     5644..5663
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5882..5986)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct 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 atgggtggag 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 atggggcggg cacgagatgc catcctggac gctcttgaaa acttgtcagg ggatgaactc
     1021 aaaaagtcca agatgaagct gctgacagtg caactgcgag aaggctatgg gcgcatccca
     1081 cgcggggccc tgctgcagat ggacgccata gatctcactg acaaacttgt cagctactat
     1141 ctggagtcgt atggcttgga gctcacaatg actgtgctta gagacatggg cttacaggag
     1201 ctggctgagc agctgcaaac gactaaagaa gagtctggag ctgtggcagc tgcagccagt
     1261 gtccctgctc agagtacagc cagaacagga cactttgtgg accagcacag gcaagcactc
     1321 attgccaggg tcacagaagt ggacggagtg ctggatgctt tgcatggcag tgtgctgact
     1381 gaaggacagt accaggcagt tcgtgcagag accaccagcc aagacaagat gaggaagctc
     1441 ttcagctttg ttccatcctg gaacctgacc tgcaaggact ccctcctcca ggccttgaag
     1501 gaaatacatc cctacttggt gatggacctg gagcagagct gactcgagca tgcatctaga
     1561 gggccctatt ctatagtgtc acctaaatgc tagagctcgc tgatcagcct cgactgtgcc
     1621 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     1681 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     1741 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     1801 caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag
     1861 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     1921 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     1981 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     2041 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     2101 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     2161 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     2221 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     2281 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg
     2341 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     2401 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     2461 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     2521 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     2581 gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     2641 ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag
     2701 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     2761 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     2821 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     2881 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     2941 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     3001 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     3061 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     3121 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     3181 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     3241 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     3301 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     3361 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     3421 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     3481 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga
     3541 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg
     3601 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     3661 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
     3721 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     3781 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
     3841 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     3901 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     3961 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4021 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     4081 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     4141 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     4201 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     4261 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     4321 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     4381 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     4441 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     4501 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     4561 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     4621 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     4681 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     4741 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     4801 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     4861 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     4921 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     4981 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     5041 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5101 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     5161 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     5221 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5281 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     5341 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     5401 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     5461 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     5521 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     5581 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     5641 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     5701 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     5761 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     5821 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     5881 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     5941 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     6001 aagtgccacc tgacgtc
//
