LOCUS       sequence-332701-        3381 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pDONR221 for Gateway shuttling. Contains the CARD8 FIIND 
            (L162M-P434, no stop codon) that cannot bind DPP9 at site I (E274R).
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 E274R
FEATURES             Location/Qualifiers
     source          1..3381
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     184..203
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     437..454
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(613..640)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(732..818)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     867..889
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     881..898
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     882..898
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    914..1013
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(1851..1950)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1967..1986)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1968..1986)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1991..2007)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1991..2007)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2109..2128)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2120..2929
                     /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(2187..2206)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      join(3076..3381,1..283)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
       61 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
      121 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
      181 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
      241 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
      301 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
      361 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
      421 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
      481 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
      541 ctggaaagcg ggcagtgagc gcaacgcaat taatacgcgt accgctagcc aggaagagtt
      601 tgtagaaacg caaaaaggcc atccgtcagg atggccttct gcttagtttg atgcctggca
      661 gtttatggcg ggcgtcctgc ccgccaccct ccgggccgtt gcttcacaac gttcaaatcc
      721 gctcccggcg gatttgtcct actcaggaga gcgttcaccg acaaacaaca gataaaacga
      781 aaggcccagt cttccgactg agcctttcgt tttatttgat gcctggcagt tccctactct
      841 cgcgttaacg ctagcatgga tgttttccca gtcacgacgt tgtaaaacga cggccagtct
      901 taagctcggg ccccaaataa tgattttatt ttgactgata gtgacctgtt cgttgcaaca
      961 aattgatgag caatgctttt ttataatgcc aactttgtac aaaaaagcag gcttcgaagg
     1021 agatagaacc atggggcctg aaggaaatgt ggatgttgag ttgattgata agagcacaaa
     1081 cagatacagc gtttggttcc ccactgctgg ctggtatctg tggtcagcca caggcctcgg
     1141 cttcctggta agggatgagg tcacagttac catcgccttc ggcagctgga gtcagcacct
     1201 ggccctggac ctgcagcacc atgaacagtg gctggtgggc ggccccttgt ttgatgtcac
     1261 tgcagagcca gaggaggctg tcgccgaaat ccacctcccc cacttcatct ccctccaagc
     1321 aggtgaggtg gacgtctcct ggtttctcgt tgcccatttt aagaatagag ggatggtcct
     1381 ggagcatcca gcccgggtgg agcctttcta tgctgtcctg gaaagcccca gcttctctct
     1441 gatgggcatc ctgctgcgga tcgccagtgg gactcgcctc tccatcccca tcacttccaa
     1501 cacattgatc tattatcacc cccaccccga agatattaag ttccacttgt accttgtccc
     1561 cagcgacgcc ttgctaacaa aggcgataga tgatgaggaa gatcgcttcc atggtgtgcg
     1621 cctgcagact tcgcccccaa tggaacccct gaactttggt tccagttata ttgtgtctaa
     1681 ttctgctaac ctgaaagtaa tgcccaagga gttgaaattg tcctacagga gccctggaga
     1741 aattcagcac ttctcaaaat tctatgctgg gcagatgaag gaacccattc aacttgagat
     1801 tactgaaaaa agacatggga ctttggtgtg ggatactgag gtgaagccag acccagcttt
     1861 cttgtacaaa gttggcatta taagaaagca ttgcttatca atttgttgca acgaacaggt
     1921 cactatcagt caaaataaaa tcattatttg ccatccagct gatatcccct atagtgagtc
     1981 gtattacatg gtcatagctg tttcctggca gctctggccc gtgtctcaaa atctctgatg
     2041 ttacattgca caagataaaa atatatcatc atgaacaata aaactgtctg cttacataaa
     2101 cagtaataca aggggtgtta tgagccatat tcaacgggaa acgtcgaggc cgcgattaaa
     2161 ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc
     2221 aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca
     2281 tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac
     2341 ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt
     2401 actcaccact gcgatccccg gaaaaacagc attccaggta ttagaagaat atcctgattc
     2461 aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt
     2521 ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat
     2581 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga
     2641 acaagtctgg aaagaaatgc ataaactttt gccattctca ccggattcag tcgtcactca
     2701 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga
     2761 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct
     2821 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc
     2881 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat cagaattggt
     2941 taattggttg taacactggc agagcattac gctgacttga cgggacggcg caagctcatg
     3001 accaaaatcc cttaacgtga gttacgcgtc gttccactga gcgtcagacc ccgtagaaaa
     3061 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     3121 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     3181 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     3241 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     3301 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     3361 atagttaccg gataaggcgc a
//
