LOCUS       sequence-214074-        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     102..121
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     205..224
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        205..223
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    224..248
                     /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             263..285
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             305..322
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             332..349
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             356..649
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             2167..2184
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2237..2255)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2251..2298
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2335..2790
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2422..2441)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2632..2653
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2882..3697)
                     /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(2882..3697)
                     /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     3605..3624
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3689..3708
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3819..4407
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4308..4327
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4561..4578
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4593..4732
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4658..4677)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4818..4840
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4834..5025)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5834..6916)
                     /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     6878..6897
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6917..6994)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa
       61 gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag
      121 gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg
      181 atcgagatct cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata
      241 acaattcccc tctagaaata attttgttta actttaagaa ggagatatac atatgggcag
      301 cagccatcat catcatcatc acggcagcgg cctggtgccg cgcggcagcg ctagcatgtc
      361 ggactcagaa gtcaatcaag aagctaagcc agaggtcaag ccagaagtca agcctgagac
      421 tcacatcaat ttaaaggtgt ccgatggatc ttcagagatc ttcttcaaga tcaaaaagac
      481 cactccttta agaaggctga tggaagcgtt cgctaaaaga cagggtaagg aaatggactc
      541 cttaagattc ttgtacgacg gtattagaat tcaagctgat cagacccctg aagatttgga
      601 catggaggat aacgatatta ttgaggctca cagagaacag attggtggta tggggtcggc
      661 ctttgagcgg gtagtccgga gagtggtcca ggagctggac catggtgggg agttcatccc
      721 tgtgaccagc ctgcagagct ccactggctt ccagccctac tgcctggtgg ttaggaagcc
      781 ctcaagctca tggttctgga aaccccgtta taagtgtgtc aacctgtcta tcaaggacat
      841 cctggagccg gatgccgcgg aaccagacgt gcagcgtggc aggagcttcc acttctacga
      901 tgccatggat gggcagatac agggcagcgt ggagctggca gccccaggac aggcaaagat
      961 cgcaggcggg gccgcggtgt ctgacagctc cagcacctca atgaatgtgt actcgctgag
     1021 tgtggaccct aacacctggc agactctgct ccatgagagg cacctgcggc agccagaaca
     1081 caaagtcctg cagcagctgc gcagccgcgg ggacaacgtg tacgtggtga ctgaggtgct
     1141 acagacacag aaggaggtgg aagtcacgcg cacccacaag cgggagggct cgggccggtt
     1201 ttccctgccc ggagccacgt gcttgcaggg tgagggccag ggccatctga gccagaagaa
     1261 gacggtcacc atcccctcag gcagcaccct cgcattccgg gtggcccagc tggttattga
     1321 ctctgacttg gacgtccttc tcttcccgga taagaagcag aggaccttcc agccacccgc
     1381 gacaggccac aagcgttcca cgagcgaagg cgcctggcca cagctgccct ctggcctctc
     1441 catgatgagg tgcctccaca acttcctgac agatggggtc cctgcggagg gggcgttcac
     1501 tgaagacttc cagggcctac gggcagaggt ggagaccatc tccaaggaac tggagctttt
     1561 ggacagagag ctgtgccagc tgctgctgga gggcctggag ggggtgctgc gggaccagct
     1621 ggccctgcga gccttggagg aggcgctgga gcagggccag agccttgggc cggtggagcc
     1681 cctggacggt ccagcaggtg ctgtcctgga gtgcctggtg ttgtcctccg gaatgctggt
     1741 gccggaactc gctatccctg ttgtctacct gctgggggca ctgaccatgc tgagtgaaac
     1801 gcagcacaag ctgctggcgg aggcgctgga gtcgcagacc ctgttggggc cgctcgagct
     1861 ggtgggcagc ctcttggagc agagtgcccc gtggcaggag cgcagcacca tgtccctgcc
     1921 ccccgggctc ctggggaaca gctggggcga aggagcaccg gcctgggtct tgctggacga
     1981 gtgtggccta gagctggggg aggacactcc ccacgtgtgc tgggagccgc aggcccaggg
     2041 ccgcatgtgt gcactctacg cctccctggc actgctatca ggactgagcc aggagcccca
     2101 ctagagacaa gcttaggtat ttattcggcg caaagtgcgt cgggtgatgc tgccaactta
     2161 gtcgagcacc accaccacca ccactgagat ccggctgcta acaaagcccg aaaggaagct
     2221 gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg
     2281 gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa tgggacgcgc
     2341 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     2401 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     2461 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     2521 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     2581 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     2641 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     2701 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     2761 attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg
     2821 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgaattaatt
     2881 cttagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa
     2941 taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc
     3001 ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac
     3061 ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga
     3121 ctgaatccgg tgagaatggc aaaagtttat gcatttcttt ccagacttgt tcaacaggcc
     3181 agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt
     3241 gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg
     3301 aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat
     3361 attcttctaa tacctggaat gctgttttcc cggggatcgc agtggtgagt aaccatgcat
     3421 catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt
     3481 ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa
     3541 acaactctgg cgcatcgggc ttcccataca atcgatagat tgtcgcacct gattgcccga
     3601 cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg
     3661 gcctagagca agacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta
     3721 tgtaagcaga cagttttatt gttcatgacc aaaatccctt aacgtgagtt ttcgttccac
     3781 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     3841 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     3901 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     3961 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     4021 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     4081 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     4141 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     4201 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     4261 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     4321 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     4381 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     4441 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     4501 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     4561 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     4621 ctgtgcggta tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat
     4681 agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca
     4741 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     4801 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     4861 acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc
     4921 tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct
     4981 gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta
     5041 agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat
     5101 acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc
     5161 ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa
     5221 tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat
     5281 aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac
     5341 cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc
     5401 gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct
     5461 caacgacagg agcacgatca tgcgcacccg tggggccgcc atgccggcga taatggcctg
     5521 cttctcgccg aaacgtttgg tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa
     5581 gattccgaat accgcaagcg acaggccgat catcgtcgcg ctccagcgaa agcggtcctc
     5641 gccgaaaatg acccagagcg ctgccggcac ctgtcctacg agttgcatga taaagaagac
     5701 agtcataagt gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt
     5761 gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct aacttacatt
     5821 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     5881 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc
     5941 ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc tgagagagtt
     6001 gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta
     6061 acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc gagatatccg
     6121 caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt
     6181 tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa
     6241 aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag
     6301 tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg
     6361 ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac
     6421 cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata
     6481 acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat
     6541 agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg
     6601 cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc
     6661 gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa
     6721 cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat
     6781 tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct
     6841 ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata
     6901 acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca
     6961 taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc tcccttatgc
     7021 gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca
     7081 aggaatggtg catgcaag
//
