LOCUS       sequence-479189-        7486 bp    DNA     circular SYN 22-JUL-2025
DEFINITION  Expression of N-terminal FLAG tagged human gasdermin B.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7486)
  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 7486)
  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-hGSDMB
FEATURES             Location/Qualifiers
     source          1..7486
                     /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(4901..4924)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(4968..5171)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4997..5017)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5172..5551
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(5712..5729)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5712..5728)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5721..5743)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(5869..6324)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6006..6027)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6218..6237
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6351..6455
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6456..6524,6525..7316)
                     /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(6674..6693)
                     /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 ggaagagaca gaggtaggcc
     3661 cctcagccag ctccagcagg atagagacaa caacatacag cgcacagaga attcgtgcct
     3721 cagggtcata gtccatgtca ggaggactgc tggccagctc atcccagttc tgctccatga
     3781 cagatttcac ctggtccttc aacagaggaa gggtcccctt ctccagggcc tcagccacaa
     3841 actgctgctc ttcagacagc tctagcaggg catccaggaa gtccagaatg gcttttgcac
     3901 gcgcttctac caagacccca gcagcattaa aaaggctgct taggagaggc ttgtctgggt
     3961 cctccatgtg tagctccccg gaaatcagga cctcagatac tctttgctct agatcctgcc
     4021 gaatatcctc cttgccgagg cacttagcga gggagtttag cacatctttt ctcttctcct
     4081 ctgtcaggtc cttgaggaca ctctccatgt cctccaactt ctccttcatg tttctggaat
     4141 cctccgaacc caaagacttt cctaaacagg atgaagcacc atccttctct tctggaaagg
     4201 attttgtttt gcccctgaaa tgaatattca tcgtctcctt gttggggaag acaagctgct
     4261 ttactcgata gctcaggacc cgatttgggg ggatggtcac ttccctttgg cccttgtgtt
     4321 tatagctgag atggccctga gagatctggc tccaaaattt atattgccgg tcgcttttca
     4381 gggtttcctc ctttaccgtc tccagagttt ctgtcaccag atacaggttt tctctcgtat
     4441 taattgatcg gaatgaaaag ggtagttccc tcttcagctt cctgttttca agggtagcca
     4501 gatactgctg ggatatccgg ttctccgata tcttgatttt ctgatggtgg aagccctgga
     4561 aactgcctga aattgttatt tctttgggta atctcactat caactctccc gttgagtcta
     4621 cattatccag aatttgaaac tcagcctttt gaccttggag cccagaatcc agttcatcta
     4681 accacttgtc cccatctgtg tccagaatgt ccatcagggt gaggcctgtt gtgtagtgcc
     4741 ggcatccaaa gaaagttctc ttctccccca ccagatggaa gcagcggaat ctatcagcat
     4801 caacaaggct tctaacggca atcatatccc ctccagcatc catctcctta actacaattc
     4861 ttgtgatttc ctcaaatacg ctgaacattg cggccgcaag cttgtcgtca tcgtctttgt
     4921 agtccatggt agatcgatct gaattaattc tgacggttca ctaaacgagc tctgcttata
     4981 tagacctccc accgtacacg cctaccgccc atttgcgtca acggggcggg gttattacga
     5041 cattttggaa agtcccgttg attttggtgc caaaacaaac tcccattgac gtcaatgggg
     5101 tggagacttg gaaatccccg tgagtcaaac cgctatccac gcccattggt gtactgccaa
     5161 aaccgcatca ccatggtaat agcgatgact aatacgtaga tgtactgcca agtaggaaag
     5221 tcccgtaagg tcatgtactg ggcataatgc caggcgggcc atttaccgtc attgacgtca
     5281 atagggggcg gacttggcat atgatacact tgatgtactg ccaagtgggc agtttaccgt
     5341 aaatactcca cccattgacg tcaatggaaa gtccctattg gcgttactat gggaacatac
     5401 gtcattattg acgtcaatgg gcgggggtcg ttgggcggtc agccaggcgg gccatttacc
     5461 gtaagttatg taacgcggaa ctccatatat gggctatgaa ctaatgaccc cgtaattgat
     5521 tactattaat aactagtcaa taatcaatgt caacatggcg gtcatattgg acatgagcca
     5581 atataaatgt acatattatg atatagatac aacgtatgca atggccaata gccaatattg
     5641 atttatgcta tataaccaat gactaatatg gctaattgcc aatattgatt caatgtatag
     5701 atcagcttgg cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     5761 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     5821 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggaa attgtaagcg
     5881 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
     5941 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
     6001 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
     6061 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
     6121 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
     6181 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
     6241 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
     6301 ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     6361 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     6421 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     6481 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     6541 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     6601 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     6661 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     6721 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     6781 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     6841 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     6901 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6961 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     7021 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     7081 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     7141 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     7201 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     7261 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     7321 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     7381 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     7441 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttc
//
