LOCUS       sequence-425994-        7773 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human BTRC. C-terminal NanoLuc 
            luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7773)
  AUTHORS   Ackloo S, Li F, Szewczyk M, Seitova A, Loppnau P, Zeng H, Xu J, 
            Ahmad S, Arnautova YA, Baghaie AJ, Beldar S, Bolotokova A, Centrella
            PA, Chau I, Clark MA, Cuozzo JW, Dehghani-Tafti S, Disch JS, Dong A,
            Dumas A, Feng JA, Ghiabi P, Gibson E, Gilmer J, Goldman B, Green SR,
            Guie MA, Guilinger JP, Harms N, Herasymenko O, Houliston S, 
            Hutchinson A, Kearnes S, Keefe AD, Kimani SW, Kramer T, Kutera M, 
            Kwak HA, Lento C, Li Y, Liu J, Loup J, Machado RAC, Mulhern CJ, 
            Perveen S, Righetto GL, Riley P, Shrestha S, Sigel EA, Silva M, 
            Sintchak MD, Slakman BL, Taylor RD, Thompson J, Torng W, 
            Underkoffler C, von Rechenberg M, Walsh RT, Watson I, Wilson DJ, 
            Wolf E, Yadav M, Yazdi AK, Zhang J, Zhang Y, Santhakumar V, Edwards 
            AM, Barsyte-Lovejoy D, Schapira M, Brown PJ, Halabelian L, 
            Arrowsmith CH
  TITLE     A Target Class Ligandability Evaluation of WD40 Repeat-Containing 
            Proteins.
  JOURNAL   J Med Chem. 2025 Jan 23;68(2):1092-1112. doi: 
            10.1021/acs.jmedchem.4c02010. Epub 2024 Nov 4.
  PUBMED    39495097
REFERENCE   2  (bases 1 to 7773)
  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: 
            "10.1021/acs.jmedchem.4c02010"; journalName: "J Med Chem"; date: 
            "2025-01-23- 23"; volume: "68"; issue: "2"; pages: "1092-1112"
COMMENT     Sequence Label: pNLF1-C_BTRC:1-605
FEATURES             Location/Qualifiers
     source          1..7773
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1033..1052
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1033..1051
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1079..1088
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2948..3460
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(3536..3554)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3591..3712
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3602..3621)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3656..3675
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3957..4314
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3957..4148
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3980..4000)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4165..4300
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4345..5382
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             5361..5369
                     /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)"
     polyA_signal    5406..5454
                     /note="synthetic polyadenylation signal"
     promoter        5610..5714
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5715..5783,5784..6575)
                     /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     complement(5933..5952)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6733..7321)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6813..6832)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    7437..7720
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt attaatacga ctcactatag ggctagcaaa gcgatcgctt ccgaattcgc
     1081 caccatggac ccggccgagg cggtgctgca agagaaggca ctcaagttta tgtgctctat
     1141 gcccaggtct ctgtggctgg gctgctccag cctggcggac agcatgcctt cgctgcgatg
     1201 cctgtataac ccagggactg gcgcactcac agctttccag aattcctcag agagagaaga
     1261 ctgtaataat ggcgaacccc ctaggaagat aataccagag aagaattcac ttagacagac
     1321 atacaacagc tgtgccagac tctgcttaaa ccaagaaaca gtatgtttag caagcactgc
     1381 tatgaagact gagaattgtg tggccaaaac aaaacttgcc aatggcactt ccagtatgat
     1441 tgtgcccaag caacggaaac tctcagcaag ctatgaaaag gaaaaggaac tgtgtgtcaa
     1501 atactttgag cagtggtcag agtcagatca agtggaattt gtggaacatc ttatatccca
     1561 aatgtgtcat taccaacatg ggcacataaa ctcgtatctt aaacctatgt tgcagagaga
     1621 tttcataact gctctgccag ctcggggatt ggatcatatt gctgagaaca ttctgtcata
     1681 cctggatgcc aaatcactat gtgctgctga acttgtgtgc aaggaatggt accgagtgac
     1741 ctctgatggc atgctgtgga agaagcttat cgagagaatg gtcaggacag attctctgtg
     1801 gagaggcctg gcagaacgaa gaggatgggg acagtattta ttcaaaaaca aacctcctga
     1861 cgggaatgct cctcccaact ctttttatag agcactttat cctaaaatta tacaagacat
     1921 tgagacaata gaatctaatt ggagatgtgg aagacatagt ttacagagaa ttcactgccg
     1981 aagtgaaaca agcaaaggag tttactgttt acagtatgat gatcagaaaa tagtaagcgg
     2041 ccttcgagac aacacaatca agatctggga taaaaacaca ttggaatgca agcgaattct
     2101 cacaggccat acaggttcag tcctctgtct ccagtatgat gagagagtga tcataacagg
     2161 atcatcggat tccacggtca gagtgtggga tgtaaataca ggtgaaatgc taaacacgtt
     2221 gattcaccat tgtgaagcag ttctgcactt gcgtttcaat aatggcatga tggtgacctg
     2281 ctccaaagat cgttccattg ctgtatggga tatggcctcc ccaactgaca ttaccctccg
     2341 gagggtgctg gtcggacacc gagctgctgt caatgttgta gactttgatg acaagtacat
     2401 tgtttctgca tctggggata gaactataaa ggtatggaac acaagtactt gtgaatttgt
     2461 aaggacctta aatggacaca aacgaggcat tgcctgtttg cagtacaggg acaggctggt
     2521 agtgagtggc tcatctgaca acactatcag attatgggac atagaatgtg gtgcatgttt
     2581 acgagtgtta gaaggccatg aggaattggt gcgttgtatt cgatttgata acaagaggat
     2641 agtcagtggg gcctatgatg gaaaaattaa agtgtgggat cttgtggctg ctttggaccc
     2701 ccgtgctcct gcagggacac tctgtctacg gacccttgtg gagcattccg gaagagtttt
     2761 tcgactacag tttgatgaat tccagattgt cagtagttca catgatgaca caatcctcat
     2821 ctgggacttc ctaaatgatc cagctgccca agctgaaccc ccccgttccc cttctcgaac
     2881 atacacctac atctccagat ctagatttgg gcccaattcc tgcaggcgag ctctcggctc
     2941 gagcggcgtc ttcacactcg aagatttcgt tggggactgg cgacagacag ccggctacaa
     3001 cctggaccaa gtccttgaac agggaggtgt gtccagtttg tttcagaatc tcggggtgtc
     3061 cgtaactccg atccaaagga ttgtcctgag cggtgaaaat gggctgaaga tcgacatcca
     3121 tgtcatcatc ccgtatgaag gtctgagcgg cgaccaaatg ggccagatcg aaaaaatttt
     3181 taaggtggtg taccctgtgg atgatcatca ctttaaggtg atcctgcact atggcacact
     3241 ggtaatcgac ggggttacgc cgaacatgat cgactatttc ggacggccgt atgaaggcat
     3301 cgccgtgttc gacggcaaaa agatcactgt aacagggacc ctgtggaacg gcaacaaaat
     3361 tatcgacgag cgcctgatca accccgacgg ctccctgctg ttccgagtaa ccatcaacgg
     3421 agtgaccggc tggcggctgt gcgaacgcat tctggcgtaa ggccgcgact ctagggcatg
     3481 caagctgatc cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct
     3541 gagcaataac tagcataacc ccttggggcg gccgcttcga gcagacatga taagatacat
     3601 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     3661 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa
     3721 caattgcatt cattttatgt ttcaggttca gggggagatg tgggaggttt ttttaagcaa
     3781 gtaaaacctc tacaaatgtg gtaaaatcga attttaacaa aatattaacg cttacaattt
     3841 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgcggatc
     3901 tgcgcagcac catggcctga aataacctct gaaagaggaa cttggttagg taccttctga
     3961 ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     4021 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     4081 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     4141 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     4201 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     4261 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     4321 gattcttctg acactagcgc caccatgaag aagcccgaac tcaccgctac cagcgttgaa
     4381 aaatttctca tcgagaagtt cgacagtgtg agcgacctga tgcagttgtc ggagggcgaa
     4441 gagagccgag ccttcagctt cgatgtcggc ggacgcggct atgtactgcg ggtgaatagc
     4501 tgcgctgatg gcttctacaa agaccgctac gtgtaccgcc acttcgccag cgctgcacta
     4561 cccatccccg aagtgttgga catcggcgag ttcagcgaga gcctgacata ctgcatcagt
     4621 agacgcgccc aaggcgttac tctccaagac ctccccgaaa cagagctgcc tgctgtgtta
     4681 cagcctgtcg ccgaagctat ggatgctatt gccgccgccg acctcagtca aaccagcggc
     4741 ttcggcccat tcgggcccca aggcatcggc cagtacacaa cctggcggga tttcatttgc
     4801 gccattgctg atccccatgt ctaccactgg cagaccgtga tggacgacac cgtgtccgcc
     4861 agcgtagctc aagccctgga cgaactgatg ctgtgggccg aagactgtcc cgaggtgcgc
     4921 cacctcgtcc atgccgactt cggcagcaac aacgtcctga ccgacaacgg ccgcatcacc
     4981 gccgtaatcg actggtccga agctatgttc ggggacagtc agtacgaggt ggccaacatc
     5041 ttcttctggc ggccctggct ggcttgcatg gagcagcaga ctcgctactt cgagcgccgg
     5101 catcccgagc tggccggcag ccctcgtctg cgagcctaca tgctgcgcat cggcctggat
     5161 cagctctacc agagcctcgt ggacggcaac ttcgacgatg ctgcctgggc tcaaggccgc
     5221 tgcgatgcca tcgtccgcag cggggccggc accgtcggtc gcacacaaat cgctcgccgg
     5281 agcgcagccg tatggaccga cggctgcgtc gaggtgctgg ccgacagcgg caaccgccgg
     5341 cccagtacac gaccgcgcgc taaggaggta ggtcgagttt agactctagc catcacgatg
     5401 gccgcaataa aatatcttta ttttcattac atctgtgtgt tggttttttg tgtgaatcga
     5461 tagcgataag gatcctcttt gcgcttgcgt tttcccttgt ccagatagcc cagtagctga
     5521 cattcatccg gggtcagcac cgtttctgcg gactggcttt ctacgtaatg gtttcttaga
     5581 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     5641 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5701 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5761 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     5821 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     5881 agagttttcg ccccgaagaa cgttttccaa tgatgagcac tttcaaagtt ctgctatgtg
     5941 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     6001 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     6061 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     6121 ttctgacaac tatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     6181 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     6241 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     6301 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     6361 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     6421 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     6481 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     6541 ctgagatagg tgcctcactg attaagcatt ggtaattcga aatgaccgac caagcgacgc
     6601 ccaaccggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     6661 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     6721 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     6781 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     6841 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     6901 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     6961 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     7021 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     7081 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     7141 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     7201 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     7261 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatttca
     7321 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     7381 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt tatagtccgg
     7441 aaatacagga acgcacgctg gatggccctt cgctgggatg gtgaaaccat gaaaaatggc
     7501 agcttcagtg gattaagtgg gggtaatgtg gcctgtaccc tctggttgca taggtattca
     7561 tacggttaaa atttatcagg cgcgattgcg gcagtttttc gggtggtttg ttgccatttt
     7621 tacctgtctg ctgccgtgat cgcgctgaac gcgttttagc ggtgcgtaca attaagggat
     7681 tatggtaaat ccacttactg tctgccctcg tagccatcga gataaaccgc agtactccgg
     7741 ccacgatgcg tccggcgtag aggatcgaga tct
//
