LOCUS       sequence-479200-        7573 bp    DNA     circular SYN 22-JUL-2025
DEFINITION  Expression of N-terminal FLAG tagged human gasdermin A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7573)
  AUTHORS   LaRock DL, Johnson AF, Wilde S, Sands JS, Monteiro MP, LaRock CN
  TITLE     Group A Streptococcus induces GSDMA-dependent pyroptosis in 
            keratinocytes.
  JOURNAL   Nature. 2022 May;605(7910):527-531. doi: 10.1038/s41586-022-04717-x.
            Epub 2022 May 11.
  PUBMED    35545676
REFERENCE   2  (bases 1 to 7573)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 22, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41586-022-04717-x"; journalName: "Nature"; date: 
            "2022-05"; volume: "605"; issue: "7910"; pages: "527-531"
COMMENT     Sequence Label: pCMV-Flag-hGSDMA
FEATURES             Location/Qualifiers
     source          1..7573
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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)."
     primer_bind     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1005..1024
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1005..1023
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    1029..1163
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1053..1072)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1107..1126
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1815..2609)
                     /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(1926..1945)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2536..2555
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(2663..2960)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     2958..2978
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    3002..3624
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     3406..3425
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     CDS             complement(4988..5011)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(5055..5258)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5084..5104)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5259..5638
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(5799..5816)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5799..5815)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5808..5830)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(5956..6411)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6093..6114)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6305..6324
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6438..6542
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6543..6611,6612..7403)
                     /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     complement(6761..6780)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga catgattacg aatttaatac gactcactat
     1021 agggaattga tccagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     1081 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     1141 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     1201 agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtatggctg
     1261 attatgatcg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttccagc
     1321 ttggcgagtg ggtggtgagc agctcgccgt cgcgcagcag gcgccagcgc tccatccacg
     1381 cagtgacctc aggcggaaaa cgggaagaca cactcatgag atgcctgcaa gcaattcgtt
     1441 ctgtatcagg cgcaggagcg tcccgtccgg gtcgaccaaa gcggccatcg tgcctcccca
     1501 ctcctgcagt tcgggggcat ggatgcgcgg atagccgctg ctggtttcct ggatgccgac
     1561 ggatttgcac tgccggtaga actccgcgag gtcgtccagc ctcaggcagc agctgaacca
     1621 actcgcgagg ggatcgagcc cggggtgggc gaagaactcc agcatgagat ccccgcgctg
     1681 gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt catagaaggc
     1741 ggcggtggaa tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg tcatttcgaa
     1801 ccccagagtc ccgctcagaa gaactcgtca agaaggcgat agaaggcgat gcgctgcgaa
     1861 tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc gccaagctct
     1921 tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac acccagccgg
     1981 ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg caagcaggca
     2041 tcgccatggg tcacgacgag atcctcgccg tcgggcatgc gcgccttgag cctggcgaac
     2101 agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc gacaagaccg
     2161 gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc gaatgggcag
     2221 gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga tactttctcg
     2281 gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa tagcagccag
     2341 tcccttcccg cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc cgtcgtggcc
     2401 agccacgata gccgcgctgc ctcgtcctgc agttcattca gggcaccgga caggtcggtc
     2461 ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga acacggcggc atcagagcag
     2521 ccgattgtct gttgtgccca gtcatagccg aatagcctct ccacccaagc ggccggagaa
     2581 cctgcgtgca atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt ctcttgatca
     2641 gatcccaagc tgatcaattc ctccaaaaaa gcctcctcac tacttctgga atagctcaga
     2701 ggccgaggcg gcctcggcct ctgcataaat aaaaaaaatt agtcagccat ggggcggaga
     2761 atgggcggaa ctgggcggag ttaggggcgg gatgggcgga gttaggggcg ggactatggt
     2821 tgctgactaa ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt
     2881 tccacacctg gttgctgact aattgagatg catgctttgc atacttctgc ctgctgggga
     2941 gcctggggac tttccacacc ctaactgaca cacattccac agccaagctg gccgcgtacc
     3001 caattcaaca ggcatctact gagtggaccc aacgcatgag aggacagtgc caagcaagca
     3061 actcaaatgt cccaccggtt gggcatggcc aggtagccta tgctgtgtct ggacgtcctc
     3121 ctgctggtat agttatttta aaatcagaag gacagggaag ggagcagtgg ttcacgcctg
     3181 taatcccagc aatttgggag gccaaggtgg gtagatcacc tgagattagg agttggagac
     3241 cagcctggcc aatatggtga aaccccgtct ctaccaaaaa aacaaaaatt agctgagcct
     3301 ggtcatgcat gcctggaatc ccaacaactc gggaggctga ggcaggagaa tcgcttgaac
     3361 ccaggaggcg gagattgcag tgagccaaga ttgtgccact gcactccagc ttggttccca
     3421 atagaccccg caggccctac aggttgtctt cccaacttgc cccttgctcc ataccacccc
     3481 cctccacccc ataatattat agaaggacac ctagtcagac aaaatgatgc aacttaattt
     3541 tattaggaca aggctggtgg gcactggagt ggcaacttcc agggccagga gaggcactgg
     3601 ggaggggtca cagggatgcc acccgggatc ctctagatta ggaggccttg gtaagctgct
     3661 gaaggagaga gaggcctgca taaagagcac agaggcgagg gagggtgtct gggtcccaca
     3721 tatattgggg gcccgatctc tgcacttcca ggccactcag cccgactaga gcctccgtga
     3781 gggtcagctc ctcgtcccca agggaggaga gcagctcagg ttgcaggggg aaaacaccct
     3841 ctttatccag caggaagttc tgttccatcg tgctctccac cagctttagc tgcacgggta
     3901 ggatcttttt ctccatggat ttcaccagca gcttctgttg ggcttcactt agctctgtta
     3961 gggctccaac gaaatagagg atggcgccca cggcctcctt tgatagcagg acatcttttg
     4021 ggagtgcctc cagggtcact tcatgtccct tgtctagagc cccttcaagt gcgagctcaa
     4081 ggtcttgtag ctcctttttc ttccctagaa gtttgctgag ggagctgagc agggagcttt
     4141 gccctactcg gctcagcttc tccacttgtt gggtctctct ctgaacttct tcttttagtg
     4201 tcctgaagcc ttcgtgtacg tccccaacat cagatgcctg gataaggatg accttctcct
     4261 ctcctgactt ttctccagga gggaaggttt gcatgttatc attgcagata tgtggaatat
     4321 cccactcatc tttgcctttg accatcagct gtctcactcg aaaggccagg acgcagccct
     4381 tggggatggt tacagcctcc ttgtgattta tggatccctg tagccccaat ggggcgaaga
     4441 aggggaggga gaagcaggcc tctgccttgc cggctcgctc cagtgtgacc tcctgcaccg
     4501 tctccaccac ctccatcacc acatacaggt tctccccttg atcttgcatc tccttcagga
     4561 atgggtggtc tgctgccagc ttcctctcct gcacggtctc cagggccttg ggagccacac
     4621 tgagtgtctg gacctccaga gtgctgttct gcgagagccc tgccgttccc ttcaccttca
     4681 ccgtctttgg tacatccaca tctccctcca ctcgggtgtc cagcatattc ttaaagccaa
     4741 aattcccagt gtctgttggg tctgaaggtg agctgccggg ctcaagcaca tccagcagcg
     4801 tgtagtcggt gcggacgtac cgggcccccc agaagagcgt gctcttcctc ttcctcagca
     4861 ccaggcagaa gggatggaag cgcttgaagt cgatgaggct gtcaagtggt gtcaggtccc
     4921 ctcgagggtt tagctgtctg gccagggccc gggtgacatt ttcaaacatg gtcattgcgg
     4981 ccgcaagctt gtcgtcatcg tctttgtagt ccatggtaga tcgatctgaa ttaattctga
     5041 cggttcacta aacgagctct gcttatatag acctcccacc gtacacgcct accgcccatt
     5101 tgcgtcaacg gggcggggtt attacgacat tttggaaagt cccgttgatt ttggtgccaa
     5161 aacaaactcc cattgacgtc aatggggtgg agacttggaa atccccgtga gtcaaaccgc
     5221 tatccacgcc cattggtgta ctgccaaaac cgcatcacca tggtaatagc gatgactaat
     5281 acgtagatgt actgccaagt aggaaagtcc cgtaaggtca tgtactgggc ataatgccag
     5341 gcgggccatt taccgtcatt gacgtcaata gggggcggac ttggcatatg atacacttga
     5401 tgtactgcca agtgggcagt ttaccgtaaa tactccaccc attgacgtca atggaaagtc
     5461 cctattggcg ttactatggg aacatacgtc attattgacg tcaatgggcg ggggtcgttg
     5521 ggcggtcagc caggcgggcc atttaccgta agttatgtaa cgcggaactc catatatggg
     5581 ctatgaacta atgaccccgt aattgattac tattaataac tagtcaataa tcaatgtcaa
     5641 catggcggtc atattggaca tgagccaata taaatgtaca tattatgata tagatacaac
     5701 gtatgcaatg gccaatagcc aatattgatt tatgctatat aaccaatgac taatatggct
     5761 aattgccaat attgattcaa tgtatagatc agcttggcac tggccgtcgt tttacaacgt
     5821 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     5881 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     5941 ctgaatggcg aatggaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     6001 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     6061 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     6121 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     6181 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     6241 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     6301 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     6361 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     6421 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     6481 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     6541 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     6601 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     6661 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     6721 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     6781 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     6841 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     6901 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     6961 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     7021 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     7081 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     7141 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     7201 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     7261 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     7321 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     7381 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     7441 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     7501 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     7561 tcaaaggatc ttc
//
