LOCUS       sequence-209480-        7098 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express human GSDMD in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7098)
  AUTHORS   Kambara H, Liu F, Zhang X, Liu P, Bajrami B, Teng Y, Zhao L, Zhou S,
            Yu H, Zhou W, Silberstein LE, Cheng T, Han M, Xu Y, Luo HR
  TITLE     Gasdermin D Exerts Anti-inflammatory Effects by Promoting Neutrophil
            Death.
  JOURNAL   Cell Rep. 2018 Mar 13;22(11):2924-2936. doi: 
            10.1016/j.celrep.2018.02.067.
  PUBMED    29539421
REFERENCE   2  (bases 1 to 7098)
  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: 
            "10.1016/j.celrep.2018.02"; journalName: "Cell Rep"; date: 
            "2018-03-13- 13"; volume: "22"; issue: "11"; pages: "2924-2936"
COMMENT     Sequence Label: pET-SUMO-hGSDMD
FEATURES             Location/Qualifiers
     source          1..7098
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     106..125
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     209..228
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        209..227
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    228..252
                     /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)."
     RBS             267..289
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             309..326
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             336..353
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             360..653
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             2171..2188
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2241..2259)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2255..2302
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2339..2794
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2426..2445)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2636..2657
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2886..3701)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2886..3701)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3609..3628
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3693..3712
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3823..4411
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4312..4331
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4565..4582
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4597..4736
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4662..4681)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4822..4844
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4838..5029)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5838..6920)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     6882..6901
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6921..6998)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
       61 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
      121 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
      181 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
      241 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat atacatatgg
      301 gcagcagcca tcatcatcat catcacggca gcggcctggt gccgcgcggc agcgctagca
      361 tgtcggactc agaagtcaat caagaagcta agccagaggt caagccagaa gtcaagcctg
      421 agactcacat caatttaaag gtgtccgatg gatcttcaga gatcttcttc aagatcaaaa
      481 agaccactcc tttaagaagg ctgatggaag cgttcgctaa aagacagggt aaggaaatgg
      541 actccttaag attcttgtac gacggtatta gaattcaagc tgatcagacc cctgaagatt
      601 tggacatgga ggataacgat attattgagg ctcacagaga acagattggt ggtatggggt
      661 cggcctttga gcgggtagtc cggagagtgg tccaggagct ggaccatggt ggggagttca
      721 tccctgtgac cagcctgcag agctccactg gcttccagcc ctactgcctg gtggttagga
      781 agccctcaag ctcatggttc tggaaacccc gttataagtg tgtcaacctg tctatcaagg
      841 acatcctgga gccggatgcc gcggaaccag acgtgcagcg tggcaggagc ttccacttct
      901 acgatgccat ggatgggcag atacagggca gcgtggagct ggcagcccca ggacaggcaa
      961 agatcgcagg cggggccgcg gtgtctgaca gctccagcac ctcaatgaat gtgtactcgc
     1021 tgagtgtgga ccctaacacc tggcagactc tgctccatga gaggcacctg cggcagccag
     1081 aacacaaagt cctgcagcag ctgcgcagcc gcggggacaa cgtgtacgtg gtgactgagg
     1141 tgctgcagac acagaaggag gtggaagtca cgcgcaccca caagcgggag ggctcgggcc
     1201 ggttttccct gcccggagcc acgtgcttgc agggtgaggg ccagggccat ctgagccaga
     1261 agaagacggt caccatcccc tcaggcagca ccctcgcatt ccgggtggcc cagctggtta
     1321 ttgactctga cttggacgtc cttctcttcc cggataagaa gcagaggacc ttccagccac
     1381 ccgcgacagg ccacaagcgt tccacgagcg aaggcgcctg gccacagctg ccctctggcc
     1441 tctccatgat gaggtgcctc cacaacttcc tgacagatgg ggtccctgcg gagggggcgt
     1501 tcactgaaga cttccagggc ctacgggcag aggtggagac catctccaag gaactggagc
     1561 ttttggacag agagctgtgc cagctgctgc tggagggcct ggagggggtg ctgcgggacc
     1621 agctggccct gcgagccttg gaggaggcgc tggagcaggg ccagagcctt gggccggtgg
     1681 agcccctgga cggtccagca ggtgctgtcc tggagtgcct ggtgttgtcc tccggaatgc
     1741 tggtgccgga actcgctatc cctgttgtct acctgctggg ggcactgacc atgctgagtg
     1801 aaacgcagca caagctgctg gcggaggcgc tggagtcgca gaccctgttg gggccgctcg
     1861 agctggtggg cagcctcttg gagcagagtg ccccgtggca ggagcgcagc accatgtccc
     1921 tgccccccgg gctcctgggg aacagctggg gcgaaggagc accggcctgg gtcttgctgg
     1981 acgagtgtgg cctagagctg ggggaggaca ctccccacgt gtgctgggag ccgcaggccc
     2041 agggccgcat gtgtgcactc tacgcctccc tggcactgct atcaggactg agccaggagc
     2101 cccactagag acaagcttag gtatttattc ggcgcaaagt gcgtcgggtg atgctgccaa
     2161 cttagtcgag caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga
     2221 agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa
     2281 acgggtcttg aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac
     2341 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     2401 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     2461 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     2521 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     2581 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     2641 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     2701 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     2761 gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc
     2821 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgaatt
     2881 aattcttaga aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta
     2941 tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag
     3001 ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata
     3061 caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg
     3121 acgactgaat ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca
     3181 ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt
     3241 gattgcgcct gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga
     3301 atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca
     3361 ggatattctt ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat
     3421 gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc
     3481 cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc
     3541 agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc
     3601 ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat
     3661 cgcggcctag agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg
     3721 tttatgtaag cagacagttt tattgttcat gaccaaaatc ccttaacgtg agttttcgtt
     3781 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     3841 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     3901 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     3961 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4021 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4081 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4141 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4201 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4261 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4321 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4381 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4441 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4501 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     4561 gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac
     4621 gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg ctctgatgcc
     4681 gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc
     4741 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     4801 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     4861 cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga
     4921 tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc
     4981 ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg
     5041 tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca
     5101 cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac
     5161 tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg
     5221 ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga
     5281 acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga
     5341 agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc
     5401 gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg
     5461 tcctcaacga caggagcacg atcatgcgca cccgtggggc cgccatgccg gcgataatgg
     5521 cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt
     5581 gcaagattcc gaataccgca agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt
     5641 cctcgccgaa aatgacccag agcgctgccg gcacctgtcc tacgagttgc atgataaaga
     5701 agacagtcat aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg
     5761 ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta
     5821 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     5881 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt
     5941 tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag
     6001 agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg
     6061 gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata
     6121 tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga
     6181 tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt
     6241 tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg
     6301 cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg
     6361 cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc
     6421 gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga
     6481 aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc
     6541 ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta
     6601 caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg
     6661 gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg
     6721 gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg
     6781 taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg
     6841 gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg
     6901 tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat
     6961 gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt
     7021 atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc
     7081 cgcaaggaat ggtgcatg
//
