LOCUS       sequence-214079-        7107 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mouse GSDMD in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7107)
  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 7107)
  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-mGSDMD
FEATURES             Location/Qualifiers
     source          1..7107
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     109..128
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     212..231
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        212..230
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    231..255
                     /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             270..292
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             312..329
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             339..356
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             363..656
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             2183..2200
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2253..2271)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2267..2314
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2351..2806
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2438..2457)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2648..2669
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2898..3713)
                     /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(2898..3713)
                     /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     3621..3640
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3705..3724
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3835..4423
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4324..4343
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4577..4594
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4609..4748
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4674..4693)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4834..4856
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4850..5041)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5850..6932)
                     /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     6894..6913
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6933..7010)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc
       61 gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc
      121 gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc
      181 gtagaggatc gagatctcga tcccgcgaaa ttaatacgac tcactatagg ggaattgtga
      241 gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga gatatacata
      301 tgggcagcag ccatcatcat catcatcacg gcagcggcct ggtgccgcgc ggcagcgcta
      361 gcatgtcgga ctcagaagtc aatcaagaag ctaagccaga ggtcaagcca gaagtcaagc
      421 ctgagactca catcaattta aaggtgtccg atggatcttc agagatcttc ttcaagatca
      481 aaaagaccac tcctttaaga aggctgatgg aagcgttcgc taaaagacag ggtaaggaaa
      541 tggactcctt aagattcttg tacgacggta ttagaattca agctgatcag acccctgaag
      601 atttggacat ggaggataac gatattattg aggctcacag agaacagatt ggtggtatgc
      661 catcggcctt tgagaaagtg gtcaagaatg tgatcaagga ggtaagcggc agcagaggcg
      721 atctcattcc ggtggacagc ctgcggaact ccaccagctt caggccctac tgccttctga
      781 acaggaaatt ttcaagctca aggttctgga aaccccgtta ttcatgtgtc aacctgtcaa
      841 tcaaggacat cctggagccc agtgctccag aaccagaacc ggagtgtttt ggctccttca
      901 aagtctctga tgtcgtcgat gggaacattc agggcagagt gatgttgtca ggcatgggag
      961 aagggaaaat ttctggtggg gctgcagtgt ctgacagttc cagtgcctcc atgaatgtgt
     1021 gtatactgcg tgtgactcag aagacctggg agaccatgca gcatgaaagg caccttcagc
     1081 agcctgagaa caaaatcctg caacagcttc ggagtcgtgg ggatgacctg tttgtggtga
     1141 ccgaggtgct gcagacaaag gaggaagtgc agatcactga ggtccacagc caagagggct
     1201 caggccagtt tacactgcct ggagctttat gcttgaaggg tgaaggcaag ggccaccaaa
     1261 gccggaagaa gatggtgacc attcctgcag gcagcatcct ggcattccga gtggcccaac
     1321 tgcttattgg ctctaaatgg gatatccttc tcgtctcaga tgagaaacag aggacctttg
     1381 agccctcctc aggtgacaga aaagcagtgg gccagaggca ccatggcctc aatgtgcttg
     1441 ctgcgctttg ttccatcgga aagcagctca gtctcctgtc agatgggatt gatgaggagg
     1501 aattaattga ggcggcagac ttccagggcc tgtatgctga ggtgaaggct tgctcctcag
     1561 aactggagag cttggaaatg gagttgagac aacagatact ggtgaacatc gggaagattt
     1621 tacaggacca gcccagcatg gaagccttag aggcctcact agggcagggc ctgtgcagtg
     1681 gcggccaggt ggagcctctg gacggcccag ctggctgcat ccttgagtgt ctggtgcttg
     1741 actctggaga actggtgccg gaactcgcag cccctatctt ctacctgctg ggagcactgg
     1801 ctgtgctgag tgaaacccag cagcagctgc tagctaaggc tctggagaca acggtgctgt
     1861 caaagcagct ggagttggtg aagcacgtct tggaacagag caccccgtgg caggagcaga
     1921 gttctgtgtc cctgcccacc gtgctccttg gggactgctg ggatgaaaag aatcccacct
     1981 gggtcttgct agaagaatgt ggcctaaggc tgcaggtaga atccccccag gtgcactggg
     2041 aaccaacgtc tctgatcccc acaagtgcgc tctatgcctc cctgttccta ttgtcaagtc
     2101 taggccagaa accttgttag agacaagctt aggtatttat tcggcgcaaa gtgcgtcggg
     2161 tgatgctgcc aacttagtcg agcaccacca ccaccaccac tgagatccgg ctgctaacaa
     2221 agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag cataacccct
     2281 tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggatt
     2341 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     2401 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     2461 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2521 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     2581 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     2641 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     2701 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     2761 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
     2821 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2881 cgctcatgaa ttaattctta gaaaaactca tcgagcatca aatgaaactg caatttattc
     2941 atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac
     3001 tcaccgaggc agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt
     3061 ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa
     3121 tcaccatgag tgacgactga atccggtgag aatggcaaaa gtttatgcat ttctttccag
     3181 acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg
     3241 ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa
     3301 ttacaaacag gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt
     3361 tcacctgaat caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg
     3421 gtgagtaacc atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata
     3481 aattccgtca gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct
     3541 ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc
     3601 gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg
     3661 ttggaattta atcgcggcct agagcaagac gtttcccgtt gaatatggct cataacaccc
     3721 cttgtattac tgtttatgta agcagacagt tttattgttc atgaccaaaa tcccttaacg
     3781 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     3841 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     3901 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     3961 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     4021 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     4081 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     4141 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     4201 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     4261 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4321 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     4381 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4441 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4501 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4561 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     4621 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     4681 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     4741 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4801 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4861 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     4921 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     4981 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     5041 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     5101 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     5161 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     5221 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     5281 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     5341 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     5401 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     5461 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     5521 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     5581 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     5641 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     5701 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     5761 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     5821 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     5881 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5941 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     6001 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     6061 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     6121 actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     6181 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     6241 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     6301 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     6361 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     6421 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca
     6481 gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc
     6541 tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc
     6601 accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca
     6661 cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc
     6721 agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg
     6781 cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca
     6841 gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac
     6901 tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc
     6961 gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg
     7021 acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt
     7081 gagcaccgcc gccgcaagga atggtgc
//
