LOCUS       sequence-332700-        3381 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pDONR221 for Gateway shuttling. Contains the CARD8 FIIND 
            (L162M-P434, no stop codon) with an interface III mutation (R394E).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3381)
  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 3381)
  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 FIIND R394E
FEATURES             Location/Qualifiers
     source          1..3381
                     /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(1385..1484)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1501..1520)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1502..1520)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1525..1541)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1525..1541)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1643..1662)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1654..2463
                     /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(1721..1740)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2610..3198
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3099..3118
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3352..3369
                     /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 aaggagatag aaccatgggg cctgaaggaa atgtggatgt tgagttgatt
      601 gataagagca caaacagata cagcgtttgg ttccccactg ctggctggta tctgtggtca
      661 gccacaggcc tcggcttcct ggtaagggat gaggtcacag ttaccatcgc cttcggcagc
      721 tggagtcagc acctggccct ggacctgcag caccatgaac agtggctggt gggcggcccc
      781 ttgtttgatg tcactgcaga gccagaggag gctgtcgccg aaatccacct cccccacttc
      841 atctccctcc aagcaggtga ggtggacgtc tcctggtttc tcgttgccca ttttaagaat
      901 gaagggatgg tcctggagca tccagcccgg gtggagcctt tctatgctgt cctggaaagc
      961 cccagcttct ctctgatggg catcctgctg cggatcgcca gtgggactcg cctctccatc
     1021 cccatcactt ccaacacatt gatctattat cacccccacc ccgaagatat taagttccac
     1081 ttgtaccttg tccccagcga cgccttgcta acaaaggcga tagatgatga ggaagatcgc
     1141 ttccatggtg tgcgcctgca gacttcgccc ccaatggaac ccctgaactt tggttccagt
     1201 tatattgtgt ctaattctgc taacctgaaa gtaatgccca aggagttgaa attgtcctac
     1261 gagagccctg gagaaattca gcacttctca aaattctatg ctgggcagat gaaggaaccc
     1321 attcaacttg agattactga aaaaagacat gggactttgg tgtgggatac tgaggtgaag
     1381 ccagacccag ctttcttgta caaagttggc attataagaa agcattgctt atcaatttgt
     1441 tgcaacgaac aggtcactat cagtcaaaat aaaatcatta tttgccatcc agctgatatc
     1501 ccctatagtg agtcgtatta catggtcata gctgtttcct ggcagctctg gcccgtgtct
     1561 caaaatctct gatgttacat tgcacaagat aaaaatatat catcatgaac aataaaactg
     1621 tctgcttaca taaacagtaa tacaaggggt gttatgagcc atattcaacg ggaaacgtcg
     1681 aggccgcgat taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat
     1741 aatgtcgggc aatcaggtgc gacaatctat cgcttgtatg ggaagcccga tgcgccagag
     1801 ttgtttctga aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga
     1861 ctaaactggc tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct
     1921 gatgatgcat ggttactcac cactgcgatc cccggaaaaa cagcattcca ggtattagaa
     1981 gaatatcctg attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg
     2041 cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg tctcgctcag
     2101 gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg attttgatga cgagcgtaat
     2161 ggctggcctg ttgaacaagt ctggaaagaa atgcataaac ttttgccatt ctcaccggat
     2221 tcagtcgtca ctcatggtga tttctcactt gataacctta tttttgacga ggggaaatta
     2281 ataggttgta ttgatgttgg acgagtcgga atcgcagacc gataccagga tcttgccatc
     2341 ctatggaact gcctcggtga gttttctcct tcattacaga aacggctttt tcaaaaatat
     2401 ggtattgata atcctgatat gaataaattg cagtttcatt tgatgctcga tgagtttttc
     2461 taatcagaat tggttaattg gttgtaacac tggcagagca ttacgctgac ttgacgggac
     2521 ggcgcaagct catgaccaaa atcccttaac gtgagttacg cgtcgttcca ctgagcgtca
     2581 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     2641 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     2701 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     2761 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     2821 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     2881 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     2941 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     3001 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     3061 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     3121 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     3181 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     3241 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     3301 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     3361 gtgagcgagg aagcggaaga g
//
