LOCUS       sequence-267548-        7066 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Gsdmd-NT mutant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7066)
  AUTHORS   Liu X, Zhang Z, Ruan J, Pan Y, Magupalli VG, Wu H, Lieberman J
  TITLE     Inflammasome-activated gasdermin D causes pyroptosis by forming 
            membrane pores.
  JOURNAL   Nature. 2016 Jul 6;535(7610):153-8. doi: 10.1038/nature18629.
  PUBMED    27383986
REFERENCE   2  (bases 1 to 7066)
  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"; journalName: 
            "Nature"; date: "2016-07-6- 6"; volume: "535"; issue: "7610"; pages:
            "153-8"
COMMENT     Sequence Label: Flag-Gsdmd-NT C39A
FEATURES             Location/Qualifiers
     source          1..7066
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     98..120
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     112..129
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     113..129
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        290..669
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        670..873
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     824..844
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             917..940
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    1797..2419
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(1996..2015)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2443..2463)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2461..2758
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2812..3606
                     /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(2866..2885)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3476..3495
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4258..4392
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4295..4314)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4349..4368
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4397..4416)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4398..4416)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4430..4446)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4430..4446)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4443..4465)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4454..4470
                     /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(4478..4484,4485..4502,4503..4508))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4523..4544
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4661..4678)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4832..5420)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4912..4931)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5591..6382,6383..6451))
                     /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     6214..6233
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6452..6556)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6583..7038
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6670..6689)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6880..6901
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
       61 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
      121 acggccagtg ccaagctgat ctatacattg aatcaatatt ggcaattagc catattagtc
      181 attggttata tagcataaat caatattggc tattggccat tgcatacgtt gtatctatat
      241 cataatatgt acatttatat tggctcatgt ccaatatgac cgccatgttg acattgatta
      301 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      361 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      421 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      481 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      541 atgccaagtc cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      601 cagtacatga ccttacggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      661 attaccatgg tgatgcggtt ttggcagtac accaatgggc gtggatagcg gtttgactca
      721 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      781 caacgggact ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat gggcggtagg
      841 cgtgtacggt gggaggtcta tataagcaga gctcgtttag tgaaccgtca gaattaattc
      901 agatcgatct accatggact acaaagacga tgacgacaag cttgcggccg caatgccatc
      961 ggcctttgag aaagtggtca agaatgtgat caaggaggta agcggcagca gaggcgatct
     1021 cattccggtg gacagcctgc ggaactccac cagcttcagg ccctacgccc ttctgaacag
     1081 gaaattttca agctcaaggt tctggaaacc ccgttattca tgtgtcaacc tgtcaatcaa
     1141 ggacatcctg gagcccagtg ctccagaacc agaaccggag tgttttggct ccttcaaagt
     1201 ctctgatgtc gtcgatggga acattcaggg cagagtgatg ttgtcaggca tgggagaagg
     1261 gaaaatttct ggtggggctg cagtgtctga cagttccagt gcctccatga atgtgtgtat
     1321 actgcgtgtg actcagaaga cctgggagac catgcagcat gaaaggcacc ttcagcagcc
     1381 tgagaacaaa atcctgcaac agcttcggag tcgtggggat gacctgtttg tggtgaccga
     1441 ggtgctgcag acaaaggagg aagtgcagat cactgaggtc cacagccaag agggctcagg
     1501 ccagtttacg ctgcctggag ctttatgctt gaagggtgaa ggcaagggcc accaaagccg
     1561 gaagaagatg gtgaccattc ctgcaggcag catcctggca ttccgagtgg cccaactgct
     1621 tattggctct aaatgggata tccttctcgt ctcagatgag aaacagagga cctttgagcc
     1681 ctcctcaggt gacagaaaag cagtgggcca gaggcaccat ggcctcaatg tgcttgctgc
     1741 gctttgttcc atcggaaagc agctcagtct cctgtcagat tagtctagag gatcccgggt
     1801 ggcatccctg tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc
     1861 ccaccagcct tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct
     1921 ataatattat ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct
     1981 gtagggcctg cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca
     2041 ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg
     2101 gattccaggc atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt
     2161 tcaccatatt ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc
     2221 ctcccaaatt gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt
     2281 taaaataact ataccagcag gaggacgtcc agacacagca taggctacct ggccatgccc
     2341 aaccggtggg acatttgagt tgcttgcttg gcactgtcct ctcatgcgtt gggtccactc
     2401 agtagatgcc tgttgaattg ggtacgcggc cagcttggct gtggaatgtg tgtcagttag
     2461 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2521 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     2581 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     2641 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     2701 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     2761 gaattgatca gcttgggatc tgatcaagag acaggatgag gatcgtttcg catgattgaa
     2821 caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac
     2881 tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg
     2941 cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag
     3001 gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt
     3061 gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg
     3121 tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg
     3181 catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga
     3241 gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag
     3301 gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat
     3361 ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt
     3421 tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg
     3481 gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt
     3541 tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc
     3601 ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac
     3661 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     3721 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccaccccg
     3781 ggctcgatcc cctcgcgagt tggttcagct gctgcctgag gctggacgac ctcgcggagt
     3841 tctaccggca gtgcaaatcc gtcggcatcc aggaaaccag cagcggctat ccgcgcatcc
     3901 atgcccccga actgcaggag tggggaggca cgatggccgc tttggtcgac ccggacggga
     3961 cgctcctgcg cctgatacag aacgaattgc ttgcaggcat ctcatgagtg tgtcttcccg
     4021 ttttccgcct gaggtcactg cgtggatgga gcgctggcgc ctgctgcgcg acggcgagct
     4081 gctcaccacc cactcgccaa gctggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     4141 taggctccgc cgatcataat cagccatacc acatttgtag aggttttact tgctttaaaa
     4201 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac
     4261 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     4321 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     4381 catgtctgga tcaattccct atagtgagtc gtattaaatt cgtaatcatg tcatagctgt
     4441 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     4501 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     4561 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     4621 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     4681 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4741 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4801 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4861 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4921 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4981 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     5041 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     5101 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     5161 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     5221 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     5281 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5341 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     5401 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     5461 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5521 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5581 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5641 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5701 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5761 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5821 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5881 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5941 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     6001 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6061 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6121 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6181 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6241 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6301 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6361 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6421 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6481 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6541 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgcgccc tgtagcggcg
     6601 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     6661 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     6721 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     6781 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     6841 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     6901 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     6961 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     7021 tattaacgct tacaatttcc attcgccatt caggctgcgc aactgt
//
