LOCUS       sequence-335588-        4161 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pDONR221 for Gateway shuttling. Contains the CARD8 T60 isoform (no 
            stop codon).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4161)
  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 4161)
  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
FEATURES             Location/Qualifiers
     source          1..4161
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(147..174)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(266..352)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     401..423
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     415..432
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     416..432
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    448..547
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(2165..2264)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2281..2300)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2282..2300)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2305..2321)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2305..2321)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2423..2442)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2434..3243
                     /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     complement(2501..2520)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3390..3978
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3879..3898
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4132..4149
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac
       61 gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatac gcgtaccgct
      121 agccaggaag agtttgtaga aacgcaaaaa ggccatccgt caggatggcc ttctgcttag
      181 tttgatgcct ggcagtttat ggcgggcgtc ctgcccgcca ccctccgggc cgttgcttca
      241 caacgttcaa atccgctccc ggcggatttg tcctactcag gagagcgttc accgacaaac
      301 aacagataaa acgaaaggcc cagtcttccg actgagcctt tcgttttatt tgatgcctgg
      361 cagttcccta ctctcgcgtt aacgctagca tggatgtttt cccagtcacg acgttgtaaa
      421 acgacggcca gtcttaagct cgggccccaa ataatgattt tattttgact gatagtgacc
      481 tgttcgttgc aacaaattga tgagcaatgc ttttttataa tgccaacttt gtacaaaaaa
      541 gcaggcttcg ccatggaaaa aaaggagtgt ccagaaaaga gtagcagcag tgaggaagag
      601 ctgccgagac gggacagtgg atccagtagg aacatagatg catccaaact cattagacta
      661 caaggatcac ggaaactgtt ggttgacaat agcatacggg aactgcaata cacaaaaact
      721 ggaatttttt ttcaggctga ggcctgtgtg acaaatgata cggtatacag ggagctaccc
      781 tgtgtttctg agaccctttg tgacatctca cattttttcc aagaagatga tgagacagag
      841 gcagagccat tattgttccg tgctgttcct gagtgtcaac tatctggggg ggacattccc
      901 agtgtatcag aagagcagga atcttcagag ggacaagatt caggagacat ttgctcagaa
      961 gagaatcaaa tagtttcctc ttatgcttct aaagtctgtt ttgagatcga agaagattat
     1021 aaaaatcgtc agtttctggg gcctgaagga aatgtggatg ttgagttgat tgataagagc
     1081 acaaacagat acagcgtttg gttccccact gctggctggt atctgtggtc agccacaggc
     1141 ctcggcttcc tggtaaggga tgaggtcaca gtgacgattg cgtttggttc ctggagtcag
     1201 cacctggccc tggacctgca gcaccatgaa cagtggctgg tgggcggccc cttgtttgat
     1261 gtcactgcag agccagagga ggctgtcgcc gaaatccacc tcccccactt catctccctc
     1321 caagcaggtg aggtggacgt ctcctggttt ctcgttgccc attttaagaa tgaagggatg
     1381 gtcctggagc atccagcccg ggtggagcct ttctatgctg tcctggaaag ccccagcttc
     1441 tctctgatgg gcatcctgct gcggatcgcc agtgggactc gcctctccat ccccatcact
     1501 tccaacacat tgatctatta tcacccccac cccgaagata ttaagttcca cttgtacctt
     1561 gtccccagcg acgccttgct aacaaaggcg atagatgatg aggaagatcg cttccatggt
     1621 gtgcgcctgc agacttcgcc cccaatggaa cccctgaact ttggttccag ttatattgtg
     1681 tctaattctg ctaacctgaa agtaatgccc aaggagttga aattgtccta caggagccct
     1741 ggagaaattc agcacttctc aaaattctat gctgggcaga tgaaggaacc cattcaactt
     1801 gagattactg aaaaaagaca tgggactttg gtgtgggata ctgaggtgaa gccagtggat
     1861 ctccagcttg tagctgcatc agcccctcct cctttctcag gtgcagcctt tgtgaaggag
     1921 aaccaccggc aactccaagc caggatgggg gacctgaaag gggtgctcga tgatctccaa
     1981 gacaatgagg ttcttactga gaatgagaag gagctggtgg agcaggaaaa gacacggcag
     2041 agcaagaatg aggccttgct gagcatggtg gagaagaaag gggacctggc cctggacgtg
     2101 ctcttcagaa gcattagtga aagggaccct tacctcgtgt cctatcttag acagcagaat
     2161 ttggacccag ctttcttgta caaagttggc attataagaa agcattgctt atcaatttgt
     2221 tgcaacgaac aggtcactat cagtcaaaat aaaatcatta tttgccatcc agctgatatc
     2281 ccctatagtg agtcgtatta catggtcata gctgtttcct ggcagctctg gcccgtgtct
     2341 caaaatctct gatgttacat tgcacaagat aaaaatatat catcatgaac aataaaactg
     2401 tctgcttaca taaacagtaa tacaaggggt gttatgagcc atattcaacg ggaaacgtcg
     2461 aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat
     2521 aatgtcgggc aatcaggtgc gacaatctat cgcttgtatg ggaagcccga tgcgccagag
     2581 ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga
     2641 ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct
     2701 gatgatgcat ggttactcac cactgcgatc cccggaaaaa cagcattcca ggtattagaa
     2761 gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg
     2821 cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg tctcgctcag
     2881 gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga cgagcgtaat
     2941 ggctggcctg ttgaacaagt ctggaaagaa atgcataaac ttttgccatt ctcaccggat
     3001 tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga ggggaaatta
     3061 ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga tcttgccatc
     3121 ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt tcaaaaatat
     3181 ggtattgata atcctgatat gaataaattg cagtttcatt tgatgctcga tgagtttttc
     3241 taatcagaat tggttaattg gttgtaacac tggcagagca ttacgctgac ttgacgggac
     3301 ggcgcaagct catgaccaaa atcccttaac gtgagttacg cgtcgttcca ctgagcgtca
     3361 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     3421 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     3481 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     3541 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     3601 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     3661 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     3721 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     3781 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     3841 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     3901 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     3961 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     4021 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     4081 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     4141 gtgagcgagg aagcggaaga g
//
