LOCUS       sequence-266223-        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 C57A
FEATURES             Location/Qualifiers
     source          1..7066
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        105..484
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        485..688
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     639..659
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             732..755
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    1612..2234
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(1811..1830)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2258..2278)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2276..2573
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2627..3421
                     /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(2681..2700)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3291..3310
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4073..4207
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4110..4129)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4164..4183
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4212..4231)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4213..4231)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4245..4261)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4245..4261)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4258..4280)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4269..4285
                     /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(4293..4299,4300..4317,4318..4323))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4338..4359
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4476..4493)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4647..5235)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4727..4746)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5406..6197,6198..6266))
                     /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     6029..6048
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6267..6371)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6398..6853
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6485..6504)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6695..6716
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6979..7001
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6993..7010
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6994..7010
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ttatatagca taaatcaata ttggctattg gccattgcat acgttgtatc tatatcataa
       61 tatgtacatt tatattggct catgtccaat atgaccgcca tgttgacatt gattattgac
      121 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
      181 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      241 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      301 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      361 aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      421 catgacctta cgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      481 catggtgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg actcacgggg
      541 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      601 ggactttcca aaatgtcgta ataaccccgc cccgttgacg caaatgggcg gtaggcgtgt
      661 acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagaatt aattcagatc
      721 gatctaccat ggactacaaa gacgatgacg acaagcttgc ggccgcaatg ccatcggcct
      781 ttgagaaagt ggtcaagaat gtgatcaagg aggtaagcgg cagcagaggc gatctcattc
      841 cggtggacag cctgcggaac tccaccagct tcaggcccta ctgccttctg aacaggaaat
      901 tttcaagctc aaggttctgg aaaccccgtt attcagctgt caacctgtca atcaaggaca
      961 tcctggagcc cagtgctcca gaaccagaac cggagtgttt tggctccttc aaagtctctg
     1021 atgtcgtcga tgggaacatt cagggcagag tgatgttgtc aggcatggga gaagggaaaa
     1081 tttctggtgg ggctgcagtg tctgacagtt ccagtgcctc catgaatgtg tgtatactgc
     1141 gtgtgactca gaagacctgg gagaccatgc agcatgaaag gcaccttcag cagcctgaga
     1201 acaaaatcct gcaacagctt cggagtcgtg gggatgacct gtttgtggtg accgaggtgc
     1261 tgcagacaaa ggaggaagtg cagatcactg aggtccacag ccaagagggc tcaggccagt
     1321 ttacgctgcc tggagcttta tgcttgaagg gtgaaggcaa gggccaccaa agccggaaga
     1381 agatggtgac cattcctgca ggcagcatcc tggcattccg agtggcccaa ctgcttattg
     1441 gctctaaatg ggatatcctt ctcgtctcag atgagaaaca gaggaccttt gagccctcct
     1501 caggtgacag aaaagcagtg ggccagaggc accatggcct caatgtgctt gctgcgcttt
     1561 gttccatcgg aaagcagctc agtctcctgt cagattagtc tagaggatcc cgggtggcat
     1621 ccctgtgacc cctccccagt gcctctcctg gccctggaag ttgccactcc agtgcccacc
     1681 agccttgtcc taataaaatt aagttgcatc attttgtctg actaggtgtc cttctataat
     1741 attatggggt ggaggggggt ggtatggagc aaggggcaag ttggtaagac aacctgtagg
     1801 gcctgcgggg tctattggga accaagctgg agtgcagtgg cacaatcttg gctcactgca
     1861 atctccgcct cctgggttca agcgattctc ctgcctcagc ctcccgagtt gttgggattc
     1921 caggcatgca tgaccaggct cagctaattt ttgttttttt ggtagagacg gggtttcacc
     1981 atattggcca ggctggtctc caactcctaa tctcaggtga tctacccacc ttggcctccc
     2041 aaattgctgg gattacaggc gtgaaccact gctcccttcc ctgtccttct gattttaaaa
     2101 taactatacc agcaggagga cgtccagaca cagcataggc tacctggcca tgcccaaccg
     2161 gtgggacatt tgagttgctt gcttggcact gtcctctcat gcgttgggtc cactcagtag
     2221 atgcctgttg aattgggtac gcggccagct tggctgtgga atgtgtgtca gttagggtgt
     2281 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     2341 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     2401 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     2461 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     2521 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggaatt
     2581 gatcagcttg ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga
     2641 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     2701 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     2761 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc
     2821 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     2881 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     2941 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     3001 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     3061 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     3121 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt
     3181 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     3241 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     3301 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     3361 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     3421 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     3481 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     3541 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccgggctc
     3601 gatcccctcg cgagttggtt cagctgctgc ctgaggctgg acgacctcgc ggagttctac
     3661 cggcagtgca aatccgtcgg catccaggaa accagcagcg gctatccgcg catccatgcc
     3721 cccgaactgc aggagtgggg aggcacgatg gccgctttgg tcgacccgga cgggacgctc
     3781 ctgcgcctga tacagaacga attgcttgca ggcatctcat gagtgtgtct tcccgttttc
     3841 cgcctgaggt cactgcgtgg atggagcgct ggcgcctgct gcgcgacggc gagctgctca
     3901 ccacccactc gccaagctgg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     3961 tccgccgatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct
     4021 cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt
     4081 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4141 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4201 ctggatcaat tccctatagt gagtcgtatt aaattcgtaa tcatgtcata gctgtttcct
     4261 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     4321 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     4381 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     4441 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4501 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     4561 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     4621 cgtaaaaagg ccgcgttgtt ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4681 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4741 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4801 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4861 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     4921 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     4981 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5041 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     5101 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5161 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     5221 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     5281 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     5341 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     5401 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     5461 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     5521 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     5581 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     5641 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     5701 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5761 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5821 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     5881 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     5941 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     6001 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     6061 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     6121 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     6181 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     6241 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     6301 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     6361 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg cgccctgtag cggcgcatta
     6421 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     6481 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     6541 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     6601 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     6661 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     6721 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     6781 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     6841 acgcttacaa tttccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc
     6901 gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt
     6961 gggtaacgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt gccaagctga
     7021 tctatacatt gaatcaatat tggcaattag ccatattagt cattgg
//
