LOCUS       sequence-266225-        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 C77A
FEATURES             Location/Qualifiers
     source          1..7066
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     109..131
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     123..140
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     124..140
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        301..680
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        681..884
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     835..855
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             928..951
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    1808..2430
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2007..2026)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2454..2474)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2472..2769
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2823..3617
                     /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(2877..2896)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3487..3506
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4269..4403
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4306..4325)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4360..4379
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4408..4427)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4409..4427)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4441..4457)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4441..4457)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4454..4476)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4465..4481
                     /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(4489..4495,4496..4513,4514..4519))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4534..4555
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4672..4689)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4843..5431)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4923..4942)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5602..6393,6394..6462))
                     /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     6225..6244
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6463..6567)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6594..7049
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6681..6700)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6891..6912
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
       61 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
      121 gttgtaaaac gacggccagt gccaagctga tctatacatt gaatcaatat tggcaattag
      181 ccatattagt cattggttat atagcataaa tcaatattgg ctattggcca ttgcatacgt
      241 tgtatctata tcataatatg tacatttata ttggctcatg tccaatatga ccgccatgtt
      301 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
      361 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      421 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      481 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      541 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      601 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      661 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      721 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      781 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      841 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      901 agaattaatt cagatcgatc taccatggac tacaaagacg atgacgacaa gcttgcggcc
      961 gcaatgccat cggcctttga gaaagtggtc aagaatgtga tcaaggaggt aagcggcagc
     1021 agaggcgatc tcattccggt ggacagcctg cggaactcca ccagcttcag gccctactgc
     1081 cttctgaaca ggaaattttc aagctcaagg ttctggaaac cccgttattc atgtgtcaac
     1141 ctgtcaatca aggacatcct ggagcccagt gctccagaac cagaaccgga ggcttttggc
     1201 tccttcaaag tctctgatgt cgtcgatggg aacattcagg gcagagtgat gttgtcaggc
     1261 atgggagaag ggaaaatttc tggtggggct gcagtgtctg acagttccag tgcctccatg
     1321 aatgtgtgta tactgcgtgt gactcagaag acctgggaga ccatgcagca tgaaaggcac
     1381 cttcagcagc ctgagaacaa aatcctgcaa cagcttcgga gtcgtgggga tgacctgttt
     1441 gtggtgaccg aggtgctgca gacaaaggag gaagtgcaga tcactgaggt ccacagccaa
     1501 gagggctcag gccagtttac gctgcctgga gctttatgct tgaagggtga aggcaagggc
     1561 caccaaagcc ggaagaagat ggtgaccatt cctgcaggca gcatcctggc attccgagtg
     1621 gcccaactgc ttattggctc taaatgggat atccttctcg tctcagatga gaaacagagg
     1681 acctttgagc cctcctcagg tgacagaaaa gcagtgggcc agaggcacca tggcctcaat
     1741 gtgcttgctg cgctttgttc catcggaaag cagctcagtc tcctgtcaga ttagtctaga
     1801 ggatcccggg tggcatccct gtgacccctc cccagtgcct ctcctggccc tggaagttgc
     1861 cactccagtg cccaccagcc ttgtcctaat aaaattaagt tgcatcattt tgtctgacta
     1921 ggtgtccttc tataatatta tggggtggag gggggtggta tggagcaagg ggcaagttgg
     1981 gaagacaacc tgtagggcct gcggggtcta ttgggaacca agctggagtg cagtggcaca
     2041 atcttggctc actgcaatct ccgcctcctg ggttcaagcg attctcctgc ctcagcctcc
     2101 cgagttgttg ggattccagg catgcatgac caggctcagc taatttttgt ttttttggta
     2161 gagacggggt ttcaccatat tggccaggct ggtctccaac tcctaatctc aggtgatcta
     2221 cccaccttgg cctcccaaat tgctgggatt acaggcgtga accactgctc ccttccctgt
     2281 ccttctgatt ttaaaataac tataccagca ggaggacgtc cagacacagc ataggctacc
     2341 tggccatgcc caaccggtgg gacatttgag ttgcttgctt ggcactgtcc tctcatgcgt
     2401 tgggtccact cagtagatgc ctgttgaatt gggtacgcgg ccagcttggc tgtggaatgt
     2461 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     2521 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     2581 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     2641 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     2701 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     2761 cttttttgga ggaattgatc agcttgggat ctgatcaaga gacaggatga ggatcgtttc
     2821 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     2881 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     2941 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     3001 tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     3061 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     3121 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     3181 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     3241 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     3301 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg
     3361 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     3421 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     3481 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     3541 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     3601 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     3661 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     3721 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     3781 cgcccacccc gggctcgatc ccctcgcgag ttggttcagc tgctgcctga ggctggacga
     3841 cctcgcggag ttctaccggc agtgcaaatc cgtcggcatc caggaaacca gcagcggcta
     3901 tccgcgcatc catgcccccg aactgcagga gtggggaggc acgatggccg ctttggtcga
     3961 cccggacggg acgctcctgc gcctgataca gaacgaattg cttgcaggca tctcatgagt
     4021 gtgtcttccc gttttccgcc tgaggtcact gcgtggatgg agcgctggcg cctgctgcgc
     4081 gacggcgagc tgctcaccac ccactcgcca agctggaacc gtaaaaaggc cgcgttgctg
     4141 gcgtttttcc ataggctccg ccgatcataa tcagccatac cacatttgta gaggttttac
     4201 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     4261 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     4321 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     4381 atgtatctta tcatgtctgg atcaattccc tatagtgagt cgtattaaat tcgtaatcat
     4441 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4501 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4561 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4621 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4681 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4741 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4801 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4861 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4921 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4981 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     5041 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     5101 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     5161 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     5221 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     5281 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     5341 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5401 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5461 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5521 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     5581 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5641 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5701 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5761 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5821 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5881 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     5941 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     6001 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     6061 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     6121 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     6181 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     6241 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     6301 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6361 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6421 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6481 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6541 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgcgcc
     6601 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     6661 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     6721 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     6781 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     6841 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     6901 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     6961 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     7021 ttttaacaaa atattaacgc ttacaatttc cattcgccat tcaggc
//
