LOCUS       sequence-426012-        7455 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human CDC20. C-terminal NanoLuc 
            luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7455)
  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 7455)
  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_CDC20:1-499
FEATURES             Location/Qualifiers
     source          1..7455
                     /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             2630..3142
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(3218..3236)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3273..3394
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3284..3303)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3338..3357
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3639..3996
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3639..3830
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3662..3682)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3847..3982
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4027..5064
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             5043..5051
                     /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    5088..5136
                     /note="synthetic polyadenylation signal"
     promoter        5292..5396
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5397..5465,5466..6257)
                     /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(5615..5634)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6415..7003)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6495..6514)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    7119..7402
                     /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 caccatggca cagttcgcgt tcgagagtga cctgcactcg ctgcttcagc tggatgcacc
     1141 catccccaat gcaccccctg cgcgctggca gcgcaaagcc aaggaagccg caggcccggc
     1201 cccctcaccc atgcgggccg ccaaccgatc ccacagcgcc ggcaggactc cgggccgaac
     1261 tcctggcaaa tccagttcca aggttcagac cactcctagc aaacctggcg gtgaccgcta
     1321 tatcccccat cgcagtgctg cccagatgga ggtggccagc ttcctcctga gcaaggagaa
     1381 ccagcctgaa aacagccaga cgcccaccaa gaaggaacat cagaaagcct gggctttgaa
     1441 cctgaacggt tttgatgtag aggaagccaa gatccttcgg ctcagtggaa aaccacaaaa
     1501 tgcgccagag ggttaccaga acagactgaa agtactctac agccaaaagg ccactcctgg
     1561 ctccagccgg aagacctgcc gttacattcc ttccctgcca gaccgtatcc tggatgcgcc
     1621 tgaaatccga aatgactatt acctgaacct tgtggattgg agttctggga atgtactggc
     1681 cgtggcactg gacaacagtg tgtacctgtg gagtgcaagc tctggtgaca tcctgcagct
     1741 tttgcaaatg gagcagcctg gggaatatat atcctctgtg gcctggatca aagagggcaa
     1801 ctacttggct gtgggcacca gcagtgctga ggtgcagcta tgggatgtgc agcagcagaa
     1861 acggcttcga aatatgacca gtcactctgc ccgagtgggc tccctaagct ggaacagcta
     1921 tatcctgtcc agtggttcac gttctggcca catccaccac catgatgttc gggtagcaga
     1981 acaccatgtg gccacactga gtggccacag ccaggaagtg tgtgggctgc gctgggcccc
     2041 agatggacga catttggcca gtggtggtaa tgataacttg gtcaatgtgt ggcctagtgc
     2101 tcctggagag ggtggctggg ttcctctgca gacattcacc cagcatcaag gggctgtcaa
     2161 ggccgtagca tggtgtccct ggcagtccaa tgtcctggca acaggagggg gcaccagtga
     2221 tcgacacatt cgcatctgga atgtgtgctc tggggcctgt ctgagtgccg tggatgccca
     2281 ttcccaggtg tgctccatcc tctggtctcc ccattacaag gagctcatct caggccatgg
     2341 ctttgcacag aaccagctag ttatttggaa gtacccaacc atggccaagg tggctgaact
     2401 caaaggtcac acatcccggg tcctgagtct gaccatgagc ccagatgggg ccacagtggc
     2461 atccgcagca gcagatgaga ccctgaggct atggcgctgt tttgagttgg accctgcgcg
     2521 gcggcgggag cgggagaagg ccagtgcagc caaaagcagc ctcatccacc aaggcatccg
     2581 ctctagattt gggcccaatt cctgcaggcg agctctcggc tcgagcggcg tcttcacact
     2641 cgaagatttc gttggggact ggcgacagac agccggctac aacctggacc aagtccttga
     2701 acagggaggt gtgtccagtt tgtttcagaa tctcggggtg tccgtaactc cgatccaaag
     2761 gattgtcctg agcggtgaaa atgggctgaa gatcgacatc catgtcatca tcccgtatga
     2821 aggtctgagc ggcgaccaaa tgggccagat cgaaaaaatt tttaaggtgg tgtaccctgt
     2881 ggatgatcat cactttaagg tgatcctgca ctatggcaca ctggtaatcg acggggttac
     2941 gccgaacatg atcgactatt tcggacggcc gtatgaaggc atcgccgtgt tcgacggcaa
     3001 aaagatcact gtaacaggga ccctgtggaa cggcaacaaa attatcgacg agcgcctgat
     3061 caaccccgac ggctccctgc tgttccgagt aaccatcaac ggagtgaccg gctggcggct
     3121 gtgcgaacgc attctggcgt aaggccgcga ctctagggca tgcaagctga tccggctgct
     3181 aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa
     3241 ccccttgggg cggccgcttc gagcagacat gataagatac attgatgagt ttggacaaac
     3301 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
     3361 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
     3421 gtttcaggtt cagggggaga tgtgggaggt ttttttaagc aagtaaaacc tctacaaatg
     3481 tggtaaaatc gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct
     3541 ccttacgcat ctgtgcggta tttcacaccg catacgcgga tctgcgcagc accatggcct
     3601 gaaataacct ctgaaagagg aacttggtta ggtaccttct gaggcggaaa gaaccagctg
     3661 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3721 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3781 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3841 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3901 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     3961 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcgattcttc tgacactagc
     4021 gccaccatga agaagcccga actcaccgct accagcgttg aaaaatttct catcgagaag
     4081 ttcgacagtg tgagcgacct gatgcagttg tcggagggcg aagagagccg agccttcagc
     4141 ttcgatgtcg gcggacgcgg ctatgtactg cgggtgaata gctgcgctga tggcttctac
     4201 aaagaccgct acgtgtaccg ccacttcgcc agcgctgcac tacccatccc cgaagtgttg
     4261 gacatcggcg agttcagcga gagcctgaca tactgcatca gtagacgcgc ccaaggcgtt
     4321 actctccaag acctccccga aacagagctg cctgctgtgt tacagcctgt cgccgaagct
     4381 atggatgcta ttgccgccgc cgacctcagt caaaccagcg gcttcggccc attcgggccc
     4441 caaggcatcg gccagtacac aacctggcgg gatttcattt gcgccattgc tgatccccat
     4501 gtctaccact ggcagaccgt gatggacgac accgtgtccg ccagcgtagc tcaagccctg
     4561 gacgaactga tgctgtgggc cgaagactgt cccgaggtgc gccacctcgt ccatgccgac
     4621 ttcggcagca acaacgtcct gaccgacaac ggccgcatca ccgccgtaat cgactggtcc
     4681 gaagctatgt tcggggacag tcagtacgag gtggccaaca tcttcttctg gcggccctgg
     4741 ctggcttgca tggagcagca gactcgctac ttcgagcgcc ggcatcccga gctggccggc
     4801 agccctcgtc tgcgagccta catgctgcgc atcggcctgg atcagctcta ccagagcctc
     4861 gtggacggca acttcgacga tgctgcctgg gctcaaggcc gctgcgatgc catcgtccgc
     4921 agcggggccg gcaccgtcgg tcgcacacaa atcgctcgcc ggagcgcagc cgtatggacc
     4981 gacggctgcg tcgaggtgct ggccgacagc ggcaaccgcc ggcccagtac acgaccgcgc
     5041 gctaaggagg taggtcgagt ttagactcta gccatcacga tggccgcaat aaaatatctt
     5101 tattttcatt acatctgtgt gttggttttt tgtgtgaatc gatagcgata aggatcctct
     5161 ttgcgcttgc gttttccctt gtccagatag cccagtagct gacattcatc cggggtcagc
     5221 accgtttctg cggactggct ttctacgtaa tggtttctta gacgtcaggt ggcacttttc
     5281 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     5341 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     5401 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     5461 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     5521 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     5581 aacgttttcc aatgatgagc actttcaaag ttctgctatg tggcgcggta ttatcccgta
     5641 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     5701 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     5761 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca actatcggag
     5821 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     5881 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     5941 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     6001 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     6061 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     6121 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     6181 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     6241 tgattaagca ttggtaattc gaaatgaccg accaagcgac gcccaaccgg tatcagctca
     6301 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     6361 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     6421 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     6481 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     6541 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     6601 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     6661 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     6721 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     6781 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     6841 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     6901 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     6961 tgtttgcaag cagcagatta cgcgcagaaa aaaaggattt caagaagatc ctttgatctt
     7021 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     7081 attatcaaaa aggatcttca cctagatcct tttatagtcc ggaaatacag gaacgcacgc
     7141 tggatggccc ttcgctggga tggtgaaacc atgaaaaatg gcagcttcag tggattaagt
     7201 gggggtaatg tggcctgtac cctctggttg cataggtatt catacggtta aaatttatca
     7261 ggcgcgattg cggcagtttt tcgggtggtt tgttgccatt tttacctgtc tgctgccgtg
     7321 atcgcgctga acgcgtttta gcggtgcgta caattaaggg attatggtaa atccacttac
     7381 tgtctgccct cgtagccatc gagataaacc gcagtactcc ggccacgatg cgtccggcgt
     7441 agaggatcga gatct
//
