LOCUS       sequence-332699-        3504 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pDONR221 for Gateway shuttling. Contains the CARD8 C-terminus fused 
            to ubiquitin (co-translationally processed).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3504)
  AUTHORS   Sharif H, Hollingsworth LR, Griswold AR, Hsiao JC, Wang Q, 
            Bachovchin DA, Wu H
  TITLE     Dipeptidyl peptidase 9 sets a threshold for CARD8 inflammasome 
            formation by sequestering its active C-terminal fragment.
  JOURNAL   Immunity. 2021 Jul 13;54(7):1392-1404.e10. doi: 
            10.1016/j.immuni.2021.04.024. Epub 2021 May 20.
  PUBMED    34019797
REFERENCE   2  (bases 1 to 3504)
  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.immuni.2021.04.024"; journalName: "Immunity"; date: 
            "2021-07-13- 13"; volume: "54"; issue: "7"; pages: "1392-1404.e10"
COMMENT     Sequence Label: pDONR221 CARD8 Ub-S297
FEATURES             Location/Qualifiers
     source          1..3504
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(18..35)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(189..777)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(269..288)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(924..1733)
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1647..1666
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1725..1744
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     1846..1862
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1846..1862
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1867..1886
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1867..1885
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1903..2002
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(2727..2954)
                     /codon_start=1
                     /product="human ubiquitin"
                     /label=ubiquitin
                     /translation="MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIF
                     AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     protein_bind    complement(2963..3062)
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3078..3095)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3078..3094)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3087..3109)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     terminator      3158..3244
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3336..3363
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
       61 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
      121 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
      181 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
      241 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
      301 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
      361 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
      421 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
      481 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
      541 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
      601 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
      661 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
      721 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
      781 gatcctttga tcttttctac ggggtctgac gctcagtgga acgacgcgta actcacgtta
      841 agggattttg gtcatgagct tgcgccgtcc cgtcaagtca gcgtaatgct ctgccagtgt
      901 tacaaccaat taaccaattc tgattagaaa aactcatcga gcatcaaatg aaactgcaat
      961 ttattcatat caggattatc aataccatat ttttgaaaaa gccgtttctg taatgaagga
     1021 gaaaactcac cgaggcagtt ccataggatg gcaagatcct ggtatcggtc tgcgattccg
     1081 actcgtccaa catcaataca acctattaat ttcccctcgt caaaaataag gttatcaagt
     1141 gagaaatcac catgagtgac gactgaatcc ggtgagaatg gcaaaagttt atgcatttct
     1201 ttccagactt gttcaacagg ccagccatta cgctcgtcat caaaatcact cgcatcaacc
     1261 aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa atacgcgatc gctgttaaaa
     1321 ggacaattac aaacaggaat cgaatgcaac cggcgcagga acactgccag cgcatcaaca
     1381 atattttcac ctgaatcagg atattcttct aatacctgga atgctgtttt tccggggatc
     1441 gcagtggtga gtaaccatgc atcatcagga gtacggataa aatgcttgat ggtcggaaga
     1501 ggcataaatt ccgtcagcca gtttagtctg accatctcat ctgtaacatc attggcaacg
     1561 ctacctttgc catgtttcag aaacaactct ggcgcatcgg gcttcccata caagcgatag
     1621 attgtcgcac ctgattgccc gacattatcg cgagcccatt tatacccata taaatcagca
     1681 tccatgttgg aatttaatcg cggcctcgac gtttcccgtt gaatatggct cataacaccc
     1741 cttgtattac tgtttatgta agcagacagt tttattgttc atgatgatat atttttatct
     1801 tgtgcaatgt aacatcagag attttgagac acgggccaga gctgccagga aacagctatg
     1861 accatgtaat acgactcact ataggggata tcagctggat ggcaaataat gattttattt
     1921 tgactgatag tgacctgttc gttgcaacaa attgataagc aatgctttct tataatgcca
     1981 actttgtaca agaaagctgg gtccaaattc tgctgtctaa gataggacac gaggtaaggg
     2041 tccctttcac taatgcttct gaagagcacg tccagggcca ggtccccttt cttctccacc
     2101 atgctcagca aggcctcatt cttgctctgc cgtgtctttt cctgctccac cagctccttc
     2161 tcattctcag taagaacctc attgtcttgg agatcatcga gcaccccttt caggtccccc
     2221 atcctggctt ggagttgccg gtggttctcc ttcacaaagg ctgcacctga gaaaggagga
     2281 ggggctgatg cagctacaag ctggagatcc actggcttca cctcagtatc ccacaccaaa
     2341 gtcccatgtc ttttttcagt aatctcaagt tgaatgggtt ccttcatctg cccagcatag
     2401 aattttgaga agtgctgaat ttctccaggg ctcctgtagg acaatttcaa ctccttgggc
     2461 attactttca ggttagcaga attagacaca atataactgg aaccaaagtt caggggttcc
     2521 attgggggcg aagtctgcag gcgcacacca tggaagcgat cttcctcatc atctatcgcc
     2581 tttgttagca aggcgtcgct ggggacaagg tacaagtgga acttaatatc ttcggggtgg
     2641 gggtgataat agatcaatgt gttggaagtg atggggatgg agaggcgagt cccactggcg
     2701 atccgcagca ggatgcccat cagagaacca cctcttagtc ttaagacaag atgtaaggtc
     2761 gactccttct gaatgttgta atcagacagc gttctaccgt cctcgagctg cttaccggca
     2821 aagatcaatc tttgttgatc aggtggaatt ccttccttgt cttgaatttt agccttaacg
     2881 ttttcgatgg tatcggatgg ttcaacttcc agagttatgg ttttaccggt taacgtcttg
     2941 acgaagatct gcatggtggc gaagcctgct tttttgtaca aagttggcat tataaaaaag
     3001 cattgctcat caatttgttg caacgaacag gtcactatca gtcaaaataa aatcattatt
     3061 tggggcccga gcttaagact ggccgtcgtt ttacaacgtc gtgactggga aaacatccat
     3121 gctagcgtta acgcgagagt agggaactgc caggcatcaa ataaaacgaa aggctcagtc
     3181 ggaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc tgagtaggac
     3241 aaatccgccg ggagcggatt tgaacgttgt gaagcaacgg cccggagggt ggcgggcagg
     3301 acgcccgcca taaactgcca ggcatcaaac taagcagaag gccatcctga cggatggcct
     3361 ttttgcgttt ctacaaactc ttcctggcta gcggtacgcg tattaattgc gttgcgctca
     3421 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     3481 gcggggagag gcggtttgcg tatt
//
