LOCUS       sequence-331464-        9141 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses kinase-dead NEK9 mutant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9141)
  AUTHORS   Yamamoto Y, Chino H, Tsukamoto S, Ode KL, Ueda HR, Mizushima N
  TITLE     NEK9 regulates primary cilia formation by acting as a selective 
            autophagy adaptor for MYH9/myosin IIA.
  JOURNAL   Nat Commun. 2021 Jun 2;12(1):3292. doi: 10.1038/s41467-021-23599-7.
  PUBMED    34078910
REFERENCE   2  (bases 1 to 9141)
  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: "Nat 
            Commun."; date: "2021-06-2"; volume: "12(1):3292. doi"; pages: " 
            10.1038/s41467-021-23599-7"
COMMENT     Sequence Label: pMRX-IPU-FLAG-NEK9T210A
FEATURES             Location/Qualifiers
     source          1..9141
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(550..1149)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(633..653)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     1130..1149
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     misc_feature    1167..1719
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    1169..1735
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1329..1348)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     1558..1575
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(2806..2817)
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             complement(4725..4790)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     regulatory      4790..4799
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    4826..5200
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     complement(4872..4888)
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     CDS             complement(5210..5626)
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(5226..5242)
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     complement(5258..5280)
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     misc_feature    5691..6048
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     enhancer        6407..6786
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(6888..6906)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6974..7078
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7079..7147,7148..7939)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7297..7316)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8103..8691
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8592..8611
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8845..8862
                     /label=L4440
                     /note="L4440 vector, forward primer"
     LTR             join(8992..9141,1..320)
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
ORIGIN
        1 gaggggttgt gggctctttt attgagctcg gggagcagaa gcgcgcgaac agaagcgaga
       61 agcgaactga ttggttagtt caaataaggc acagggtcat ttcaggtcct tggggcaccc
      121 tggaaacatc tgatggttct ctagaaactg ctgagggctg gaccgcatct ggggaccatc
      181 tgttcttggc cctgagccgg ggcaggaact gcttaccaca gatatcctgt ttggcccata
      241 ttcagctgtt ccatctgttc ttggccctga gccggggcag gaactgctta ccacagatat
      301 cctgtttggc ccatattcag ctgttccatc tgttcctgac cttgatctga acttctctat
      361 tctcagttat gtatttttcc atgccttgca aaatggcgtt acttaagcta gcttgccaaa
      421 cctacaggtg gggtctttca ttcccccctt tttctggaga ctaaataaaa tcttttattt
      481 tatcgtcgag ccatggggtc gtgcgctcct ttcggtcggg cgctgcgggt cgtggggcgg
      541 gcggtcgact caggcaccgg gcttgcgggt catgcaccag gtgcgcggtc cttcgggcac
      601 ctcgacgtcg gcggtgacgg tgaagccgag ccgctcgtag aaggggaggt tgcggggcgc
      661 ggaggtctcc aggaaggcgg gcaccccggc gcgctcggcc gcctccactc cggggagcac
      721 gacggcgctg cccagaccct tgccctggtg gtcgggcgag acgccgacgg tggccaggaa
      781 ccacgcgggc tccttgggcc ggtgcggcgc caggaggcct tccatctgtt gctgcgcggc
      841 cagccgggaa ccgctcaact cggccatgcg cgggccgatc tcggcgaaca ccgcccccgc
      901 ttcgacgctc tccggcgtgg tccagaccgc caccgcggcg ccgtcgtccg cgacccacac
      961 cttgccgatg tcgagcccga cgcgcgtgag gaagagttct tgcagctcgg tgacccgctc
     1021 gatgtggcgg tccgggtcga cggtgtggcg cgtggcgggg tagtcggcga acgcggcggc
     1081 gagggtgcgt acggcccggg ggacgtcgtc gcgggtggcg aggcgcaccg tgggcttgta
     1141 ctcggtcatg tcgacggttg tggccatatt atcatcgtgt ttttcaaagg aaaaccacgt
     1201 ccccgtggtt cggggggcct agacgttttt ttaacctcga ctaaacacat gtaaagcatg
     1261 tgcaccgagg ccccagatca gatcccatac aatggggtac cttctgggca tccttcagcc
     1321 ccttgttgaa tacgcttgag gagagccatt tgactctttc cacaactatc caactcacaa
     1381 cgtggcactg gggttgtgcc gcctttgcag gtgtatctta tacacgtggc ttttggccgc
     1441 agaggcacct gtcgccaggt ggggggttcc gctgcctgca aagggtcgct acagacgttg
     1501 tttgtcttca agaaacttcc agaggaactg cttccttcac gacattcaac agaccttgca
     1561 ttcctttggc gagaggggaa agacccctag gaatgctcgt caagaagaca gggccaggtt
     1621 tccgggccct cacattgcca aaagacggca atatggtgga aaataacata tagacaaacg
     1681 cacaccggcc ttattccaag cggcttcggc cagtaacgtt aggggggggg gcggaattta
     1741 cgtagcggcc gctctcgaga cccgaattcc tagaggctgg gtctacagga gtctgttccc
     1801 aggaggcacc aggaatctga atctaagtca ggctttggat ccatttccat ctcttccttt
     1861 gctgtctgag ttcctttgga atgcatcccc acctgctgcc ctccttctaa tttcttgttc
     1921 aacttctgca gttgggtaaa aatctggagg ttttcttgct gtagcttctg ttgttcagcc
     1981 agacacttta acaccagacc ctgcaatctc tcaacctcca cctgcagaga gctgcacgca
     2041 cacgcaggag gagtcagtgg tgttcccttc ccagcacagg ttactgcagg attcagccga
     2101 ggtgacgagg cttctacttg gggtttgtgt tccaaaggag cttcagaggc cactttgagt
     2161 ccttgcagct cttcataggg cagggtatct ttctcagatt ctgaaaacgc tgcactgaga
     2221 ggagatgggc tgtcaggcat ggggataaat tctgcatttt ccagctcctt tcgaagccag
     2281 ccaggacagg agctgctggc cccattactg ttccccatgg cctctgtggg actgattaaa
     2341 ccttccattc ctcggtctgc ttccattgtt cctcggaagc ctccacttgg gtcaggagtt
     2401 tcagattcct gctgactgtc ctcttcttca ccaccgccgc ccccgccgcc tcctcccggg
     2461 ctagagctct gaaacacagt tccaatggat aagccactgc tattggaacg gatggtctta
     2521 gaattcaata ctttctcaac gatgagaatg gtatgccatc cacggcaaga caggtccggg
     2581 acatgatgca gagatccaaa aataggccga ggccatgagg tacagatatc agagccatgt
     2641 ggtctctctg tgggggtcat tgccaggcgg ccattaccac cattgcccca ggcaaaaatg
     2701 tgattatcat cagtggcagc aatggtaaac tcatcaccgc aggagaccct gatcacttgc
     2761 ttcccaccaa ggggtccccc caacaggttg attcccagac gcttcttgta gttcccaacg
     2821 cccagctgcc cacacttgtt gcagccaaag gtcagcagcc ggcctcgctc atcaatagca
     2881 gctgtgtgag tcttgcctgg ggcaatggta cggatcttat aaaaggacaa ctgtttggcc
     2941 aaggtaaagg acgttgtgta gggaacttca tggtatgctt catggttgat aattcccgac
     3001 atgcactgat tcagacccag cttattgaat tcattgagtc cacaggccag cactttgcct
     3061 gactgggtca acagaaatgt cccatcacag ccacattgaa ctgcaacaat aatcaaggcc
     3121 ttgggaacat ccaccttttg tggtgtataa taatcctctt ctgaatccaa acccagtcgt
     3181 ccatattcgc cacagcccca agaatagact tccttgtttc gtgtcagaac caccacatga
     3241 ttatctccac aggagacctg ctccactgga ttgctgagga agaagttcag ctgcatgggt
     3301 tctagcactt cagggccagc aactttgtcc acccccatgc agccataata atctgatccg
     3361 aaggcataga gctgaccctc atcagtcaca cagacagtga aatcatcacc acatgacacc
     3421 tgacggatag ctttgccttg caacttttcc acatgctttg gctgtcgata ggaggctttg
     3481 tctccatggc ccagctgacc atggagttta gtgcctcctt gcatgttcac ccaagtgtac
     3541 agttccttct ccactgtgac cacagcaaag tgggtattcc ctgcacagac ctgccgggca
     3601 ctacagccac tcttgataac atccagtttc tggggggtgg attttccacc accccaaaca
     3661 tagacttcac tggttcgtga tgttactaca gcaatgggtg cttcagtcac agtgcttgac
     3721 cttggtctct ttgtaggtgc attaagcaga gtgacttttt cctccatctc tctcctgcgt
     3781 ttcctgagaa gagggcgatc tagaagttca tctgcagtag gtctctgctc aggatcctgg
     3841 tcaaggcacg aatgaaccat ttggatcaat tccaaagagt actggctaga gtcaacttcc
     3901 atggcccgaa ttccttgcac gatcttcaca cacaggttaa gtgggtttgt agcatcaaac
     3961 gtcctcttta aggtaagcag ttcaaaaatg acgcagccaa ctgcccagat atcagacttg
     4021 aaattgtact ttactccttg acagagctct ggagacatgt aatatggggt tcccacaagt
     4081 gcctcagcca tggaatactc agaattaagt ttctttgcta ggccataatc tccaagtttt
     4141 atcaggtttg ccttggtcag aaaaatattt aatgtcttta tatctctatg aaggattcca
     4201 gctttatgga tgcagctcac tgctgaaaca atctgaaata ggtaccacac caccatctct
     4261 tcctcaaaca acttgtcctt ctgacgaagg attttgtcat acaggttccc tccattacaa
     4321 tattccagct caatcagcag cgtggtattg tccatgaagt gattgtagta ggcaataatg
     4381 ttgtcgtgct gcagcagtgc cagaataact atctcattca aggcatcacg acgttccttc
     4441 tcagacagcc gggtcaaatc gacttccttc cacacaacca gtgagtcatc ctcggtgcgg
     4501 cggtacagcg tggcttcccc gaaggcgccg cggcccagga cgcggatggg gatgtagtgc
     4561 agttcctcct gctccgccgc gccgccgccg gctcgcggcc cctgactggc gctaggcccc
     4621 ggactcgagt ccccgcaacc cccggactcg ctcccaaagt ccgagttgat ggaatcgcag
     4681 tgtcgctcgt actcgcccag caccgaggat cccgagcctg aacccttgtc atcgtcatcc
     4741 ttgtagtcga tgtcatgatc tttataatca ccgtcatggt ctttgtagtc catggtggca
     4801 agctttatat cgatggcaga tccggcagtc tagaggatgg tccacccccg gggtcggcag
     4861 ccttcacgtg ggcggcgtgt atccaagctg cgatgccgtc tactttgagg gcggtggggg
     4921 tggtcagcag gactgtgtaa ggtcctttcc agcgaggttc taggttctta gtctggtgtc
     4981 ggcggaccca cactgtgtcg ccgactcggt aagggtgagg taccaccggt cggtccagtt
     5041 gttcttggta ggctgccgcc agaggtctcc agacttcgtg ctggactaag tagagagcct
     5101 gtaagtgagc ttggagagag gggctgttag taactcttgt catgtcaggg tcagggaagt
     5161 ttacaagggg cgggggtgcc ccatataaga tctcatatgg ccatatgggg gcgcctagag
     5221 aaggagtgag ggctggataa agggaggatc gaggcggggt cgaacgagga ggttcaaggg
     5281 ggagagacgg ggcggatgga ggaagaggag gcggaggctt agggtgtaca aagggcttga
     5341 cccagggagg ggggtcaaaa gccaaggctt cccaggtcac gatgtagggg acctggtctg
     5401 ggtgtccatg cgggccaggt gaaaagacct tgatcttaac ctgggtgatg aggtctcggt
     5461 taaaggtgcc gtctcgcggc catccgacgt taaaggttgg ccattctgca gagcagaagg
     5521 taacccaacg tctcttcttg acatctaccg actggttgtg agcgatccgc tcgacatctt
     5581 tccagtgacc taaggtcaaa cttaagggag tggtaacagt ctggcccggg cccatatttt
     5641 cagacaaata cagaaacaca gtcagacaga gacaacacag aacgatgctg cagcagacaa
     5701 gacgcgcggc gcggcttcgg tcccaaaccg aaagcaaaaa ttcagacgga ggcgggaact
     5761 gttttaggtt ctcgtctcct accagaacca catatccctc ctctaagggg ggtgcaccaa
     5821 agagtccaaa acgatcggga tttttggact caggtcgggc cacaaaaacg gcccccgaag
     5881 tccctgggac gtctcccagg gttgcggccg ggtgttccga actcgtcagt tccaccacgg
     5941 gtccgccaga tacagagcta gttagctaac tagtaccgac gcaggcgcat aaaatcagtc
     6001 atagacacta gacaatcgga cagacacaga taagttgctg gccagcttac ctcccggtgg
     6061 tgggtcggtg gtccctgggc aggggtctcc cgatcccgga cgagccccca aatgaaagac
     6121 ccccgctgac gggtagtcaa tcactcagag gagaccctcc caaggaacag cgagaccaca
     6181 agtcggatgc aactgcaaga gggtttattg gatacacggg tacccgggcg actcagtcaa
     6241 tcggaggact ggcgcgccga gtgaggggtt gtgggctctt ttattgagct cggggagcag
     6301 aagcgcgcga acagaagcga gaagcgaact gattggttag ttcaaataag gcacagggtc
     6361 atttcaggtc cttggggcac cctggaaaca tctgatggtt ctctagcatg gtaatagcga
     6421 tgactaatac gtagatgtac tgccaagtag gaaagtccca taaggtcatg tactgggcat
     6481 aatgccaggc gggccattta ccgtcattga cgtcaatagg gggcgtactt ggcatatgat
     6541 acacttgatg tactgccaag tgggcagttt accgtaaata ctccacccat tgacgtcaat
     6601 ggaaagtccc tattggcgtt actatgggaa catacgtcat tattgacgtc aatgggcggg
     6661 ggtcgttggg cggtcagcca ggcgggccat ttaccgtaag ttatgtaacg cggaactcca
     6721 tatatgggct atgaactaat gaccccgtaa ttgattacta ttaataacta gtcaataatc
     6781 aatgtcaaca tggcggtaat gttggacatg agccaatata aatgtacata ttatgatatg
     6841 gatacaacgt atgcaatggg ccaagctcct atagtgaaga cgaaagggcc tcgtgatacg
     6901 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     6961 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     7021 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     7081 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     7141 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     7201 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     7261 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     7321 tgttgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     7381 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     7441 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     7501 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     7561 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     7621 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     7681 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     7741 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     7801 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     7861 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     7921 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     7981 gcggccggcc gcaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     8041 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     8101 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     8161 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     8221 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     8281 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     8341 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     8401 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     8461 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     8521 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     8581 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     8641 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     8701 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     8761 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     8821 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
     8881 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcaacta tggccattta
     8941 atgtaaatac ttaagaaaaa aaaccaaatt aattttgata catgctgcat gtgaaagacc
     9001 cccgctgacg ggtagtcaat cactcagagg agaccctccc aaggaacagc gagaccacaa
     9061 gtcggatgca actgcaagag ggtttattgg atacacgggt acccgggcga ctcagtcaat
     9121 cggaggactg gcgccccgag t
//
