LOCUS       sequence-254507-        5729 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FLAG-tagged mouse TMEM98 in mammalian cells in a 
            doxycycline-dependent manner (C-terminal tag).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5729)
  AUTHORS   van der Wal T, Lambooij JP, van Amerongen R
  TITLE     TMEM98 is a negative regulator of FRAT mediated Wnt/ss-catenin 
            signalling.
  JOURNAL   PLoS One. 2020 Jan 21;15(1):e0227435. doi: 
            10.1371/journal.pone.0227435. eCollection 2020.
  PUBMED    31961879
REFERENCE   2  (bases 1 to 5729)
  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: "PLoS One.";
            date: "2020-01-21"; volume: "15(1):e0227435. doi"; pages: " 
            10.1371/journal.pone.0227435. eCollection 2020"
COMMENT     Sequence Label: pTRE-tight2-Tmem98-FLAG
FEATURES             Location/Qualifiers
     source          1..5729
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(165..269)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     337..355
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(393..415)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     polyA_signal    518..599
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(812..835)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(1548..1571)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(2272..2586)
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     primer_bind     complement(2275..2299)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    2346..2364
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2382..2400
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2417..2435
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2453..2471
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2489..2507
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2524..2542
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2560..2578
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        complement(join(2595..2601,2602..2619,2620..2625))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2640..2661
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2778..2795)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2901..2921)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2906..3235
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3086..3221
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3148..3167
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3302..3901
                     /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(3302..3321)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3798..3818
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4075..4196
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     4166..4185
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4274..4862)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4354..4373)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5033..5729,1..95,96..164))
                     /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     5656..5675
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
       61 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
      121 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
      181 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
      241 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
      301 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
      361 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
      421 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
      481 ggtgttggcg ggtgtcgggg ctggcttaac tctcgaggca gtgaaaaaaa tgctttattt
      541 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
      601 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt
      661 aaagcaagta aaacctctac aaatgtggta tggctgatta tgatcctcta gagatatcgt
      721 cgacaagctt atcgatgcgg ccgcgctagc acgcgtcagc tgactagagg atccccgggt
      781 accgagctcg aattctactc agctattaaa gcttgtcgtc atcgtctttg tagtccatgg
      841 ttgcaatggc cgactgttcc tgcaggaagc cctcagggtt ggggaggctt ttatctggct
      901 cagaagccag ggctgcctct cgaaggactt ccaggtgctc ctcagcggca gacagtgatt
      961 ggtcaatcca gtccaggccc ccggttagat ggcaggcgtt cctggtcact agcaccaagt
     1021 gactaacgga cagcagcagg gcggttgtcc gtgcatccag gagcttgggg tccagtggag
     1081 ggtacattga cttcacgacg tcgtccactc tggggctaat ccgtttggcc accacaatga
     1141 tgtcactgac acttgctgac gtcttcatct tggccccaga acccattgtc atggcaacga
     1201 gtttttctgt cagagtgtga caaatcttca agatggcgat gcagtgggac atgaggcccg
     1261 aggcatcttc gatccagtcc tcattctcca ggatggcctc gatgtggggg ttggtgatga
     1321 cgacatcatc cagctccagc tcggagggtt cagactgcgt ctccatagca ccaatgaggt
     1381 ccacgatggg cttggaatca taacgctgta gtaggtctcg aggccggcag taacgctgcc
     1441 tacagaccac caccaaagca gcaaatgagg ccaggaaaat ggtggccagc acacctatgg
     1501 cgacgatcac cacagtctcc atggcgaatt ctactcagct attaaagctt gtcgtcatcg
     1561 tctttgtagt ccatggttgc aatggccgac tgttcctgca ggaagccctc agggttgggg
     1621 aggcttttat ctggctcaga agccagggct gcctctcgaa ggacttccag gtgctcctca
     1681 gcggcagaca gtgattggtc aatccagtcc aggcccccgg ttagatggca ggcgttcctg
     1741 gtcactagca ccaagtgact aacggacagc agcagggcgg ttgtccgtgc atccaggagc
     1801 ttggggtcca gtggagggta cattgacttc acgacgtcgt ccactctggg gctaatccgt
     1861 ttggccacca caatgatgtc actgacactt gctgacgtct tcatcttggc cccagaaccc
     1921 attgtcatgg caacgagttt ttctgtcaga gtgtgacaaa tcttcaagat ggcgatgcag
     1981 tgggacatga ggcccgaggc atcttcgatc cagtcctcat tctccaggat ggcctcgatg
     2041 tgggggttgg tgatgacgac atcatccagc tccagctcgg agggttcaga ctgcgtctcc
     2101 atagcaccaa tgaggtccac gatgggcttg gaatcataac gctgtagtag gtctcgaggc
     2161 cggcagtaac gctgcctaca gaccaccacc aaagcagcaa atgaggccag gaaaatggtg
     2221 gccagcacac ctatggcgac gatcaccaca gtctccatgg cgaattctcc aggcgatctg
     2281 acggttcact aaacgagctc tgcttatata ggcctcccac cgtacacgcc tacctcgaca
     2341 tacgttctct atcactgata gggagtaaac tcgacatacg ttctctatca ctgataggga
     2401 taaactcgac atacgttctc tatcactgat agggagtaaa ctcgacatac gttctctatc
     2461 actgataggg agtaaactcg acatacgttc tctatcactg atagggagta aactcgacat
     2521 cgttctctat cactgatagg gagtaaactc gacatacgtt ctctatcact gatagggagt
     2581 aaactcgagc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     2641 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     2701 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     2761 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     2821 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     2881 ggaaagaaca tgtattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     2941 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     3001 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     3061 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     3121 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc
     3181 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     3241 cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag gatcgtttcg
     3301 catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt
     3361 acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga
     3421 ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga
     3481 catcggcaag gtatgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga
     3541 gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg
     3601 ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga
     3661 gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg
     3721 cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct
     3781 ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc
     3841 cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg
     3901 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     3961 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     4021 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     4081 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     4141 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     4201 gtctgtatac cgacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     4261 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4321 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4381 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4441 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4501 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4561 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4621 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4681 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4741 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4801 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4861 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4921 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4981 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     5041 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     5101 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     5161 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     5221 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     5281 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     5341 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     5401 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     5461 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5521 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5581 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5641 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5701 gaaaacgttc ttcggggcga aaactctca
//
