LOCUS       sequence-425956-        7227 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human FBXL2. C-terminal NanoLuc 
            luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7227)
  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 7227)
  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_FBXL2:1-423
FEATURES             Location/Qualifiers
     source          1..7227
                     /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             2402..2914
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(2990..3008)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3045..3166
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3056..3075)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3110..3129
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3411..3768
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3411..3602
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3434..3454)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3619..3754
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3799..4836
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4815..4823
                     /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    4860..4908
                     /note="synthetic polyadenylation signal"
     promoter        5064..5168
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5169..5237,5238..6029)
                     /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(5387..5406)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6187..6775)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6267..6286)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6891..7174
                     /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 caccatggtt ttctcaaaca atgatgaagg ccttattaac aaaaagttac ccaaagaact
     1141 tctgttaaga atattttcct tcttggatat agtaactttg tgccgatgtg cacagatttc
     1201 caaggcttgg aacatcttag ccctggatgg aagcaactgg caaagaatag atctttttaa
     1261 ctttcaaaca gatgtagagg gtcgagtggt ggaaaatatc tcgaagcgat gcggtggatt
     1321 cctgaggaag ctcagcttgc gaggctgcat tggtgttggg gattcctcct tgaagacctt
     1381 tgcacagaac tgccgaaaca ttgaacattt gaacctcaat ggatgcacaa aaatcactga
     1441 cagcacgtgt tatagcctta gcagattctg ttccaagctg aaacatctgg atctgacctc
     1501 ctgtgtgtct attacaaaca gctccttgaa ggggatcagt gagggctgcc gaaacctgga
     1561 gtacctgaac ctctcttggt gtgatcagat cacgaaggat ggcatcgagg cactggtgcg
     1621 aggttgtcga ggcctgaaag ccctgctcct gaggggctgc acacagttag aagatgaagc
     1681 tctgaaacac attcagaatt actgccatga gcttgtgagc ctcaacttgc agtcctgctc
     1741 acgtatcacg gatgaaggtg tggtgcagat atgcaggggc tgtcaccggc tacaggctct
     1801 ctgcctttcg ggttgcagca acctcacaga tgcctctctt acagccctgg gtttgaactg
     1861 tccgcgactg caaattttgg aggctgcccg atgctcccat ttgactgacg caggttttac
     1921 acttttagct cggaattgcc acgaattgga gaagatggat cttgaagaat gcatcctgat
     1981 aaccgacagc acactcatcc agctctccat tcactgtcct aaactgcaag ccctgagcct
     2041 gtcccactgt gaactcatca cagatgatgg gatcctgcac ctgagcaaca gtacctgtgg
     2101 ccatgagagg ctgcgggtac tggagttgga caactgcctc ctcatcactg atgtggccct
     2161 ggaacaccta gagaactgcc gaggcctgga gcgcctcgag ctgtacgact gccagcaggt
     2221 tacccgtgca ggcatcaagc ggatgcgggc tcagctccct catgtcaaag tccacgccta
     2281 ctttgctccc gtcaccccac cgacagcagt ggcaggaagt ggacagcgac tgtgcaggtg
     2341 ctgtgtcatt ctctctagat ttgggcccaa ttcctgcagg cgagctctcg gctcgagcgg
     2401 cgtcttcaca ctcgaagatt tcgttgggga ctggcgacag acagccggct acaacctgga
     2461 ccaagtcctt gaacagggag gtgtgtccag tttgtttcag aatctcgggg tgtccgtaac
     2521 tccgatccaa aggattgtcc tgagcggtga aaatgggctg aagatcgaca tccatgtcat
     2581 catcccgtat gaaggtctga gcggcgacca aatgggccag atcgaaaaaa tttttaaggt
     2641 ggtgtaccct gtggatgatc atcactttaa ggtgatcctg cactatggca cactggtaat
     2701 cgacggggtt acgccgaaca tgatcgacta tttcggacgg ccgtatgaag gcatcgccgt
     2761 gttcgacggc aaaaagatca ctgtaacagg gaccctgtgg aacggcaaca aaattatcga
     2821 cgagcgcctg atcaaccccg acggctccct gctgttccga gtaaccatca acggagtgac
     2881 cggctggcgg ctgtgcgaac gcattctggc gtaaggccgc gactctaggg catgcaagct
     2941 gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa
     3001 taactagcat aaccccttgg ggcggccgct tcgagcagac atgataagat acattgatga
     3061 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     3121 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     3181 cattcatttt atgtttcagg ttcaggggga gatgtgggag gtttttttaa gcaagtaaaa
     3241 cctctacaaa tgtggtaaaa tcgaatttta acaaaatatt aacgcttaca atttcctgat
     3301 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgcg gatctgcgca
     3361 gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt ctgaggcgga
     3421 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     3481 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3541 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3601 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3661 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     3721 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcgattct
     3781 tctgacacta gcgccaccat gaagaagccc gaactcaccg ctaccagcgt tgaaaaattt
     3841 ctcatcgaga agttcgacag tgtgagcgac ctgatgcagt tgtcggaggg cgaagagagc
     3901 cgagccttca gcttcgatgt cggcggacgc ggctatgtac tgcgggtgaa tagctgcgct
     3961 gatggcttct acaaagaccg ctacgtgtac cgccacttcg ccagcgctgc actacccatc
     4021 cccgaagtgt tggacatcgg cgagttcagc gagagcctga catactgcat cagtagacgc
     4081 gcccaaggcg ttactctcca agacctcccc gaaacagagc tgcctgctgt gttacagcct
     4141 gtcgccgaag ctatggatgc tattgccgcc gccgacctca gtcaaaccag cggcttcggc
     4201 ccattcgggc cccaaggcat cggccagtac acaacctggc gggatttcat ttgcgccatt
     4261 gctgatcccc atgtctacca ctggcagacc gtgatggacg acaccgtgtc cgccagcgta
     4321 gctcaagccc tggacgaact gatgctgtgg gccgaagact gtcccgaggt gcgccacctc
     4381 gtccatgccg acttcggcag caacaacgtc ctgaccgaca acggccgcat caccgccgta
     4441 atcgactggt ccgaagctat gttcggggac agtcagtacg aggtggccaa catcttcttc
     4501 tggcggccct ggctggcttg catggagcag cagactcgct acttcgagcg ccggcatccc
     4561 gagctggccg gcagccctcg tctgcgagcc tacatgctgc gcatcggcct ggatcagctc
     4621 taccagagcc tcgtggacgg caacttcgac gatgctgcct gggctcaagg ccgctgcgat
     4681 gccatcgtcc gcagcggggc cggcaccgtc ggtcgcacac aaatcgctcg ccggagcgca
     4741 gccgtatgga ccgacggctg cgtcgaggtg ctggccgaca gcggcaaccg ccggcccagt
     4801 acacgaccgc gcgctaagga ggtaggtcga gtttagactc tagccatcac gatggccgca
     4861 ataaaatatc tttattttca ttacatctgt gtgttggttt tttgtgtgaa tcgatagcga
     4921 taaggatcct ctttgcgctt gcgttttccc ttgtccagat agcccagtag ctgacattca
     4981 tccggggtca gcaccgtttc tgcggactgg ctttctacgt aatggtttct tagacgtcag
     5041 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     5101 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     5161 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     5221 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     5281 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     5341 ttcgccccga agaacgtttt ccaatgatga gcactttcaa agttctgcta tgtggcgcgg
     5401 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     5461 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     5521 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     5581 caactatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     5641 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     5701 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     5761 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     5821 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     5881 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     5941 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     6001 taggtgcctc actgattaag cattggtaat tcgaaatgac cgaccaagcg acgcccaacc
     6061 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     6121 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     6181 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     6241 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     6301 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     6361 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     6421 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     6481 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     6541 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     6601 gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc
     6661 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     6721 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ttcaagaaga
     6781 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     6841 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttatagt ccggaaatac
     6901 aggaacgcac gctggatggc ccttcgctgg gatggtgaaa ccatgaaaaa tggcagcttc
     6961 agtggattaa gtgggggtaa tgtggcctgt accctctggt tgcataggta ttcatacggt
     7021 taaaatttat caggcgcgat tgcggcagtt tttcgggtgg tttgttgcca tttttacctg
     7081 tctgctgccg tgatcgcgct gaacgcgttt tagcggtgcg tacaattaag ggattatggt
     7141 aaatccactt actgtctgcc ctcgtagcca tcgagataaa ccgcagtact ccggccacga
     7201 tgcgtccggc gtagaggatc gagatct
//
