LOCUS       sequence-425946-        7284 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human CRBN the C-terminal 
            NanoLuc luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7284)
  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 7284)
  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_CRBN:1-442
FEATURES             Location/Qualifiers
     source          1..7284
                     /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             2459..2971
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(3047..3065)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3102..3223
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3113..3132)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3167..3186
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3468..3825
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3468..3659
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3491..3511)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3676..3811
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3856..4893
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4872..4880
                     /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    4917..4965
                     /note="synthetic polyadenylation signal"
     promoter        5121..5225
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5226..5294,5295..6086)
                     /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(5444..5463)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6244..6832)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6324..6343)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6948..7231
                     /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 caccatggcc ggcgaaggag atcagcagga cgctgcgcac aacatgggca accacctgcc
     1141 gctcctgcct gcagagagtg aggaagaaga tgaaatggaa gttgaagacc aggatagtaa
     1201 agaagccaaa aaaccaaaca tcataaattt tgacaccagt ctgccgacat cacatacata
     1261 cctaggtgct gatatggaag aatttcatgg caggactttg cacgatgacg acagctgtca
     1321 ggtgattcca gttcttccac aagtgatgat gatcctgatt cccggacaga cattacctct
     1381 tcagcttttt caccctcaag aagtcagtat ggtgcggaat ttaattcaga aagatagaac
     1441 ctttgctgtt cttgcataca gcaatgtaca ggaaagggaa gcacagtttg gaacaacagc
     1501 agagatatat gcctatcgag aagaacagga ttttggaatt gagatagtga aagtgaaagc
     1561 aattggaaga caaaggttca aagtccttga gctaagaaca cagtcagatg gaatccagca
     1621 agctaaagtg caaattcttc ccgaatgtgt gttgccttca accatgtctg cagttcaatt
     1681 agaatccctc aataagtgcc agatatttcc ttcaaaacct gtctcaagag aagaccaatg
     1741 ttcatataaa tggtggcaga aataccagaa gagaaagttt cattgtgcaa atctaacttc
     1801 atggcctcgc tggctgtatt ccttatatga tgctgagacc ttaatggaca gaatcaagaa
     1861 acagctacgt gaatgggatg aaaatctaaa agatgattct cttccttcaa atccaataga
     1921 tttttcttac agagtagctg cttgtcttcc tattgatgat gtattgagaa ttcagctcct
     1981 taaaattggc agtgctatcc agcgacttcg ctgtgaatta gacattatga ataaatgtac
     2041 ttccctttgc tgtaaacaat gtcaagaaac agaaataaca accaaaaatg aaatattcag
     2101 tttatcctta tgtgggccga tggcagctta tgtgaatcct catggatatg tgcatgagac
     2161 acttactgtg tataaggctt gcaacttgaa tctgataggc cggccttcta cagaacacag
     2221 ctggtttcct gggtatgcct ggactgttgc ccagtgtaag atctgtgcaa gccatattgg
     2281 atggaagttt acggccacca aaaaagacat gtcacctcaa aaattttggg gcttaacgcg
     2341 atctgctctg ttgcccacga tcccagacac tgaagatgaa ataagtccag acaaagtaat
     2401 actttgcttg tctagatttg ggcccaattc ctgcaggcga gctctcggct cgagcggcgt
     2461 cttcacactc gaagatttcg ttggggactg gcgacagaca gccggctaca acctggacca
     2521 agtccttgaa cagggaggtg tgtccagttt gtttcagaat ctcggggtgt ccgtaactcc
     2581 gatccaaagg attgtcctga gcggtgaaaa tgggctgaag atcgacatcc atgtcatcat
     2641 cccgtatgaa ggtctgagcg gcgaccaaat gggccagatc gaaaaaattt ttaaggtggt
     2701 gtaccctgtg gatgatcatc actttaaggt gatcctgcac tatggcacac tggtaatcga
     2761 cggggttacg ccgaacatga tcgactattt cggacggccg tatgaaggca tcgccgtgtt
     2821 cgacggcaaa aagatcactg taacagggac cctgtggaac ggcaacaaaa ttatcgacga
     2881 gcgcctgatc aaccccgacg gctccctgct gttccgagta accatcaacg gagtgaccgg
     2941 ctggcggctg tgcgaacgca ttctggcgta aggccgcgac tctagggcat gcaagctgat
     3001 ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa
     3061 ctagcataac cccttggggc ggccgcttcg agcagacatg ataagataca ttgatgagtt
     3121 tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc
     3181 tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat
     3241 tcattttatg tttcaggttc agggggagat gtgggaggtt tttttaagca agtaaaacct
     3301 ctacaaatgt ggtaaaatcg aattttaaca aaatattaac gcttacaatt tcctgatgcg
     3361 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgcggat ctgcgcagca
     3421 ccatggcctg aaataacctc tgaaagagga acttggttag gtaccttctg aggcggaaag
     3481 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     3541 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     3601 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     3661 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     3721 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     3781 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cgattcttct
     3841 gacactagcg ccaccatgaa gaagcccgaa ctcaccgcta ccagcgttga aaaatttctc
     3901 atcgagaagt tcgacagtgt gagcgacctg atgcagttgt cggagggcga agagagccga
     3961 gccttcagct tcgatgtcgg cggacgcggc tatgtactgc gggtgaatag ctgcgctgat
     4021 ggcttctaca aagaccgcta cgtgtaccgc cacttcgcca gcgctgcact acccatcccc
     4081 gaagtgttgg acatcggcga gttcagcgag agcctgacat actgcatcag tagacgcgcc
     4141 caaggcgtta ctctccaaga cctccccgaa acagagctgc ctgctgtgtt acagcctgtc
     4201 gccgaagcta tggatgctat tgccgccgcc gacctcagtc aaaccagcgg cttcggccca
     4261 ttcgggcccc aaggcatcgg ccagtacaca acctggcggg atttcatttg cgccattgct
     4321 gatccccatg tctaccactg gcagaccgtg atggacgaca ccgtgtccgc cagcgtagct
     4381 caagccctgg acgaactgat gctgtgggcc gaagactgtc ccgaggtgcg ccacctcgtc
     4441 catgccgact tcggcagcaa caacgtcctg accgacaacg gccgcatcac cgccgtaatc
     4501 gactggtccg aagctatgtt cggggacagt cagtacgagg tggccaacat cttcttctgg
     4561 cggccctggc tggcttgcat ggagcagcag actcgctact tcgagcgccg gcatcccgag
     4621 ctggccggca gccctcgtct gcgagcctac atgctgcgca tcggcctgga tcagctctac
     4681 cagagcctcg tggacggcaa cttcgacgat gctgcctggg ctcaaggccg ctgcgatgcc
     4741 atcgtccgca gcggggccgg caccgtcggt cgcacacaaa tcgctcgccg gagcgcagcc
     4801 gtatggaccg acggctgcgt cgaggtgctg gccgacagcg gcaaccgccg gcccagtaca
     4861 cgaccgcgcg ctaaggaggt aggtcgagtt tagactctag ccatcacgat ggccgcaata
     4921 aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg atagcgataa
     4981 ggatcctctt tgcgcttgcg ttttcccttg tccagatagc ccagtagctg acattcatcc
     5041 ggggtcagca ccgtttctgc ggactggctt tctacgtaat ggtttcttag acgtcaggtg
     5101 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     5161 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     5221 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5281 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5341 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5401 gccccgaaga acgttttcca atgatgagca ctttcaaagt tctgctatgt ggcgcggtat
     5461 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5521 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5581 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5641 ctatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     5701 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     5761 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     5821 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     5881 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     5941 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6001 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     6061 gtgcctcact gattaagcat tggtaattcg aaatgaccga ccaagcgacg cccaaccggt
     6121 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     6181 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     6241 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     6301 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     6361 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     6421 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     6481 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     6541 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     6601 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     6661 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     6721 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     6781 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatttc aagaagatcc
     6841 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     6901 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttatagtccg gaaatacagg
     6961 aacgcacgct ggatggccct tcgctgggat ggtgaaacca tgaaaaatgg cagcttcagt
     7021 ggattaagtg ggggtaatgt ggcctgtacc ctctggttgc ataggtattc atacggttaa
     7081 aatttatcag gcgcgattgc ggcagttttt cgggtggttt gttgccattt ttacctgtct
     7141 gctgccgtga tcgcgctgaa cgcgttttag cggtgcgtac aattaaggga ttatggtaaa
     7201 tccacttact gtctgccctc gtagccatcg agataaaccg cagtactccg gccacgatgc
     7261 gtccggcgta gaggatcgag atct
//
