LOCUS       sequence-466920-        6320 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N-terminal HA-tagged NEK7 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6320)
  AUTHORS   Shi H, Wang Y, Li X, Zhan X, Tang M, Fina M, Su L, Pratt D, Bu CH, 
            Hildebrand S, Lyon S, Scott L, Quan J, Sun Q, Russell J, Arnett S, 
            Jurek P, Chen D, Kravchenko VV, Mathison JC, Moresco EM, Monson NL, 
            Ulevitch RJ, Beutler B
  TITLE     NLRP3 activation and mitosis are mutually exclusive events 
            coordinated by NEK7, a new inflammasome component.
  JOURNAL   Nat Immunol. 2016 Mar;17(3):250-8. doi: 10.1038/ni.3333. Epub 2015 
            Dec 7.
  PUBMED    26642356
REFERENCE   2  (bases 1 to 6320)
  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: "10.1038/ni.3333"; 
            journalName: "Nat Immunol"; date: "2016-03"; volume: "17"; issue: 
            "3"; pages: "250-8"
COMMENT     Sequence Label: pcDNA3-N-HA-NEK7
FEATURES             Location/Qualifiers
     source          1..6320
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             904..930
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     RBS             1187..1195
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     promoter        complement(1873..1891)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1874..1891)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1911..1928)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1917..2141
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2187..2615
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2274..2293)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2484..2505
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2624..2644)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2629..2958
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2809..2944
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2871..2890
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3025..3819
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3079..3098)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3689..3708
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3993..4114
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4030..4049)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4084..4103
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4163..4179)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4163..4179)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4176..4198)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4187..4203
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4211..4217,4218..4235,4236..4241))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4256..4277
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4394..4411)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4565..5153)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4645..4664)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5324..6115,6116..6184))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5947..5966
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6185..6289)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttaccatg
      901 ggatacccct acgacgtccc cgactacgcc ggatccatgg atgaacaatc acaaggaatg
      961 caagggccgc ccgttactca gttccagcca cagaaggcat tacggccaga tatgggctat
     1021 aatactttag ccaacttccg aatagaaaag aaaattggtc gtggacaatt tagtgaagtt
     1081 tatagagcat cctgtctctt ggatggagtg ccggtagcgt taaaaaaagt acagatattt
     1141 gatttaatgg atgccaaagc acgtgctgat tgtatcaaag aaatagacct ccttaagcaa
     1201 ctcaaccacc caaatgtaat taaatactat gcatcattca ttgaggataa tgagctgaac
     1261 atagttttgg agttagcaga tgctggtgac ctctccagaa tgataaagca ctttaagaaa
     1321 caaaagaggc taatccctga gagaaccgtt tggaaatact tcgttcagct ctgcagtgca
     1381 ctggaccaca tgcattctcg aagagtcatg cacagagata taaaaccagc taatgtgttc
     1441 attacagcca ctggggtagt aaaactcgga gaccttgggc ttggtcggtt tttcagctcc
     1501 aaaaccacag ccgcacattc tttagtgggt acaccttact acatgtctcc agagagaata
     1561 catgaaaatg gatacaactt caagtctgac atctggtctc ttggctgtct gctatatgag
     1621 atggctgcac tgcagagtcc tttctacggc gacaagatga acttgtattc tctgtgtaag
     1681 aagatagagc agtgtgacta cccgcctctc ccgtcagatc actattcgga ggagctacga
     1741 cagctagtta atatatgcat caacccagat ccagagaagc gacccgacat cgcctatgtt
     1801 tatgatgtgg caaagaggat gcatgcatgt accgcaagca cctaactcga gcatgcatct
     1861 agagggccct attctatagt gtcacctaaa tgctagagct cgctgatcag cctcgactgt
     1921 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     1981 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     2041 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     2101 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     2161 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     2221 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2281 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2341 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2401 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2461 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2521 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     2581 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     2641 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     2701 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2761 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     2821 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     2881 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     2941 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     3001 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     3061 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     3121 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     3181 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     3241 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     3301 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     3361 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     3421 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     3481 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     3541 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     3601 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     3661 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     3721 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     3781 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     3841 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     3901 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     3961 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     4021 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4081 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     4141 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4201 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4261 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4321 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4381 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4441 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4501 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4561 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4621 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4681 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4741 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4801 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4861 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4921 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4981 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     5041 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5101 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5161 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     5221 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     5281 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5341 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5401 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5461 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5521 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     5581 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     5641 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     5701 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     5761 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     5821 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     5881 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     5941 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6001 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6061 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6121 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6181 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6241 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6301 gaaaagtgcc acctgacgtc
//
