LOCUS       sequence-432155-        5849 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway shuttling vector for cloning N-terminal NLRP3(Delta-PYD, 
            I125M-END) tags (contains a stop codon).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5849)
  AUTHORS   Hollingsworth LR, Veeraraghavan P, Paulo JA, Harper JW
  TITLE     Spatiotemporal proteomic profiling of cellular responses to NLRP3 
            agonists
  JOURNAL   bioRxiv 2024.04.19.590338
REFERENCE   2  (bases 1 to 5849)
  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"; journalName: "bioRxiv 
            2024.04.19.590338"
COMMENT     Sequence Label: pDONR221 msNLRP3(Delta-PYD, I125M-END) (STOP)
FEATURES             Location/Qualifiers
     source          1..5849
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     489..508
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     742..759
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(918..945)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(1037..1123)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     1172..1194
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1186..1203
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1187..1203
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1219..1318
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(4067..4166)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4183..4202)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4184..4202)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4207..4223)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4207..4223)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             complement(4809..5624)
                     /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(4809..5624)
                     /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"
     rep_origin      join(5849,1..588)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
       61 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
      121 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
      181 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
      241 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
      301 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
      361 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
      421 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
      481 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
      541 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
      601 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
      661 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
      721 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata
      781 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
      841 cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc tagccaggaa
      901 gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta gtttgatgcc
      961 tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc acaacgttca
     1021 aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa caacagataa
     1081 aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg gcagttccct
     1141 actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa aacgacggcc
     1201 agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac ctgttcgttg
     1261 caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa agcaggcttc
     1321 gaaggagata gaaccatgtg taaaaagaag aaagattact gtaagatgta cagacgacat
     1381 gtgagaagca ggttctactc tatcaaggac aggaacgcgc gtctaggtga gagtgtggac
     1441 ctcaacagtc gctacacgca gctccaactg gtcaaggagc atccaagcaa gcaggagcgg
     1501 gagcatgaac tcctgaccat cggccggact aaaatgcggg acagccccat gagttccctt
     1561 aagctggagc tgctgtttga gcccgaggac gggcactcgg agcctgtgca cacagtggtg
     1621 ttccagggag cagcaggcat cgggaaaacc atcctagcca ggaagattat gttggactgg
     1681 gcactgggaa agctcttcaa agacaaattt gactatttgt tctttatcca ctgccgagag
     1741 gtgagcctca ggacgccaag gagtctagca gacctgattg tcagctgctg gcctgaccca
     1801 aacccaccag tgtgcaagat cctgcgcaag ccttccagga tcctcttcct catggatggc
     1861 tttgatgagc tacaaggggc ctttgacgag cacattgggg aggtctgcac agactggcaa
     1921 aaggctgtgc ggggagacat tctgctaagc agcctcatcc gaaagaaact gctgcccaag
     1981 gcctctctgc tcataacgac gaggccggta gccttggaga aactgcagca tctcctggac
     2041 cacccccgcc atgtggagat cctaggtttc tctgaggcca aaaggaagga gtatttcttt
     2101 aagtatttct ccaacgagct gcaggcccgg gaggccttca ggctgatcca agagaatgag
     2161 gtcctcttta ccatgtgctt catccccctg gtctgctgga ttgtgtgcac ggggctaaag
     2221 caacagatgg agaccgggaa gagcctggcc cagacctcca agaccactac ggccgtctac
     2281 gtcttcttcc tttccagcct gctgcaatcc cgggggggca ttgaggagca tctcttctct
     2341 gactacctac aggggctctg ttcactggct gcggatggaa tttggaacca gaaaatccta
     2401 tttgaggagt gtgatctgcg gaagcacggc ctgcagaaga ctgacgtctc cgctttcctg
     2461 aggatgaacg tgttccagaa ggaagtggac tgcgagagat tctacagctt cagccacatg
     2521 actttccagg agttcttcgc tgctatgtac tatttgctgg aagaggaggc agagggggag
     2581 accgtgagga aaggaccagg aggttgttca gatcttctga accgagacgt gaaggtccta
     2641 ctagaaaatt acggcaagtt tgaaaaaggc tatctgattt ttgttgtccg attcctcttt
     2701 ggccttgtaa accaggagag aacctcttat ttggagaaga aactaagttg caagatctct
     2761 cagcaagtca gactggaact actgaagtgg attgaagtga aagccaaggc caagaagctg
     2821 cagtggcagc ccagccaact ggaactgttc tactgcctgt acgagatgca ggaggaagac
     2881 tttgtgcaga gtgccatgga ccactttccc aaaattgaga tcaacctctc taccagaatg
     2941 gaccacgtgg tttcctcctt ttgtattaag aactgtcata gggtcaaaac gctttccctg
     3001 ggtttttttc acaactcgcc caaggaggaa gaagaagaga ggagaggagg tcgacccttg
     3061 gaccaggttc agtgtgtttt cccagacact catgttgcct gttcttccag actggtgaac
     3121 tgctgcctca cttctagctt ctgccgtggt ctcttctcaa gtctaagcac caaccggagc
     3181 ctcactgaac tggacctcag tgacaatact ctgggagacc cgggcatgag ggtgctgtgt
     3241 gaggcactcc agcacccagg ctgtaacatt cagagactgt ggttggggcg ctgcggactg
     3301 tcccatcaat gctgcttcga catctcctct gtcctgagca gcagccagaa gctggtggag
     3361 ctggacctca gtgacaatgc cctgggggac tttggaatca gattgctgtg tgtgggactg
     3421 aagcacctgc tctgcaacct ccagaaactg tggttggtga gctgctgtct cacatccgcg
     3481 tgttgtcagg atctcgcatt ggttctgagc tccaaccatt ctctgaccag actgtacatt
     3541 ggagaaaatg ccttgggaga ctcaggagtc caagttttgt gtgaaaagat gaaggaccca
     3601 cagtgtaact tgcagaagct ggggttggtg aattccggcc ttacttcaat ctgttgttca
     3661 gctctgacct ctgtgctcaa aaccaaccag aacttcacac acctctatct acgaagcaat
     3721 gcccttggag acacaggact caggctcctc tgtgaggggc ttctgcaccc ggactgtaaa
     3781 ctacagatgc tggaattaga caactgcagc ctcacctcac acagctgctg gaatctctcc
     3841 acaattctga cccacaacca cagccttcgg aagctgaacc tgggcaacaa tgatcttggc
     3901 gatctgtgcg tggtgaccct ctgtgaggtg ctgaaacagc agggctgcct cctgcagagc
     3961 ctacagttgg gtgaaatgta cttaaatcgt gaaacaaaac gtgccttaga agcgctccag
     4021 gaagaaaagc ctgagctgac tatagtcttc gagatttcct ggtaggaccc agctttcttg
     4081 tacaaagttg gcattataag aaagcattgc ttatcaattt gttgcaacga acaggtcact
     4141 atcagtcaaa ataaaatcat tatttgccat ccagctgata tcccctatag tgagtcgtat
     4201 tacatggtca tagctgtttc ctggcagctc tggcccgtgt ctcaaaatct ctgatgttac
     4261 attgcacaag ataaaaatat atcatcatga acaataaaac tgtctgctta cataaacagt
     4321 aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cgaggccgcg attaaattcc
     4381 aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt
     4441 gcgacaatct atcgcttgta tgggaagccc gatgcgccag agttgtttct gaaacatggc
     4501 aaaggtagcg ttgccaatga tgttacagat gagatggtca gactaaactg gctgacggaa
     4561 tttatgcctc ttccgaccat caccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4621 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4681 tctacggggt ctgacgctca gtggaacgac gcgtaactca cgttaaggga ttttggtcat
     4741 gagcttgcgc cgtcccgtca agtcagcgta atgctctgcc agtgttacaa ccaattaacc
     4801 aattctgatt agaaaaactc atcgagcatc aaatgaaact gcaatttatt catatcagga
     4861 ttatcaatac catatttttg aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg
     4921 cagttccata ggatggcaag atcctggtat cggtctgcga ttccgactcg tccaacatca
     4981 atacaaccta ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa atcaccatga
     5041 gtgacgactg aatccggtga gaatggcaaa agtttatgca tttctttcca gacttgttca
     5101 acaggccagc cattacgctc gtcatcaaaa tcactcgcat caaccaaacc gttattcatt
     5161 cgtgattgcg cctgagcgag acgaaatacg cgatcgctgt taaaaggaca attacaaaca
     5221 ggaatcgaat gcaaccggcg caggaacact gccagcgcat caacaatatt ttcacctgaa
     5281 tcaggatatt cttctaatac ctggaatgct gtttttccgg ggatcgcagt ggtgagtaac
     5341 catgcatcat caggagtacg gataaaatgc ttgatggtcg gaagaggcat aaattccgtc
     5401 agccagttta gtctgaccat ctcatctgta acatcattgg caacgctacc tttgccatgt
     5461 ttcagaaaca actctggcgc atcgggcttc ccatacaatc gatagattgt cgcacctgat
     5521 tgcccgacat tatcgcgagc ccatttatac ccatataaat cagcatccat gttggaattt
     5581 aatcgcggcc tggagcaaga cgtttcccgt tgaatatggc tcataacacc ccttgtatta
     5641 ctgtttatgt aagcagacag ttttattgtt catgatgata tatttttatc ttgtgcaatg
     5701 taacatcaga gattttgaga cacaacgtgg ctttgttgaa taaatcgaac ttttgctgag
     5761 ttgaaggatc agtcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     5821 accccgtaga aaagatcaaa ggatcttct
//
