LOCUS       sequence-423361-        7194 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human CCNE1. C-terminal NanoLuc 
            luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7194)
  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 7194)
  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_CCNE1:1-410
FEATURES             Location/Qualifiers
     source          1..7194
                     /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"
     CDS             2369..2881
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(2957..2975)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3012..3133
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3023..3042)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3077..3096
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3378..3735
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3378..3569
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3401..3421)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3586..3721
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3766..4803
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4782..4790
                     /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    4827..4875
                     /note="synthetic polyadenylation signal"
     promoter        5031..5135
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5136..5204,5205..5996)
                     /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"
     rep_origin      complement(6154..6742)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6234..6253)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6858..7141
                     /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 caccatgccg agggagcgca gggagcggga tgcgaaggag cgggacacca tgaaggagga
     1141 cggcggcgcg gagttctcgg ctcgctccag gaagaggaag gcaaacgtga ccgttttttt
     1201 gcaggatcca gatgaagaaa tggccaaaat cgacaggacg gcgagggacc agtgtgggag
     1261 ccagccttgg gacaataatg cagtctgtgc agacccctgc tccctgatcc ccacacctga
     1321 caaagaagat gatgaccggg tttacccaaa ctcaacgtgc aagcctcgga ttattgcacc
     1381 atccagaggc tccccgctgc ctgtactgag ctgggcaaat agagaggaag tctggaaaat
     1441 catgttaaac aaggaaaaga catacttaag ggatcagcac tttcttgagc aacaccctct
     1501 tctgcagcca aaaatgcgag caattcttct ggattggtta atggaggtgt gtgaagtcta
     1561 taaacttcac agggagacct tttacttggc acaagatttc tttgaccggt atatggcgac
     1621 acaagaaaat gttgtaaaaa ctcttttaca gcttattggg atttcatctt tatttattgc
     1681 agccaaactt gaggaaatct atcctccaaa gttgcaccag tttgcgtatg tgacagatgg
     1741 agcttgttca ggagatgaaa ttctcaccat ggaattaatg attatgaagg cccttaagtg
     1801 gcgtttaagt cccctgacta ttgtgtcctg gctgaatgta tacatgcagg ttgcatatct
     1861 aaatgactta catgaagtgc tactgccgca gtatccccag caaatcttta tacagattgc
     1921 agagctgttg gatctctgtg tcctggatgt tgactgcctt gaatttcctt atggtatact
     1981 tgctgcttcg gccttgtatc atttctcgtc atctgaattg atgcaaaagg tttcagggta
     2041 tcagtggtgc gacatagaga actgtgtcaa gtggatggtt ccatttgcca tggttataag
     2101 ggagacgggg agctcaaaac tgaagcactt caggggcgtc gctgatgaag atgcacacaa
     2161 catacagacc cacagagaca gcttggattt gctggacaaa gcccgagcaa agaaagccat
     2221 gttgtctgaa caaaataggg cttctcctct ccccagtggg ctcctcaccc cgccacagag
     2281 cggtaagaag cagagcagcg ggccggaaat ggcgcggata tctagatttg ggcccaattc
     2341 ctgcaggcga gctctcggct cgagcggcgt cttcacactc gaagatttcg ttggggactg
     2401 gcgacagaca gccggctaca acctggacca agtccttgaa cagggaggtg tgtccagttt
     2461 gtttcagaat ctcggggtgt ccgtaactcc gatccaaagg attgtcctga gcggtgaaaa
     2521 tgggctgaag atcgacatcc atgtcatcat cccgtatgaa ggtctgagcg gcgaccaaat
     2581 gggccagatc gaaaaaattt ttaaggtggt gtaccctgtg gatgatcatc actttaaggt
     2641 gatcctgcac tatggcacac tggtaatcga cggggttacg ccgaacatga tcgactattt
     2701 cggacggccg tatgaaggca tcgccgtgtt cgacggcaaa aagatcactg taacagggac
     2761 cctgtggaac ggcaacaaaa ttatcgacga gcgcctgatc aaccccgacg gctccctgct
     2821 gttccgagta accatcaacg gagtgaccgg ctggcggctg tgcgaacgca ttctggcgta
     2881 aggccgcgac tctagggcat gcaagctgat ccggctgcta acaaagcccg aaaggaagct
     2941 gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc ggccgcttcg
     3001 agcagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa
     3061 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
     3121 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggagat
     3181 gtgggaggtt tttttaagca agtaaaacct ctacaaatgt ggtaaaatcg aattttaaca
     3241 aaatattaac gcttacaatt tcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     3301 ttcacaccgc atacgcggat ctgcgcagca ccatggcctg aaataacctc tgaaagagga
     3361 acttggttag gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg
     3421 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3481 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3541 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     3601 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     3661 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     3721 ctaggctttt gcaaaaagct cgattcttct gacactagcg ccaccatgaa gaagcccgaa
     3781 ctcaccgcta ccagcgttga aaaatttctc atcgagaagt tcgacagtgt gagcgacctg
     3841 atgcagttgt cggagggcga agagagccga gccttcagct tcgatgtcgg cggacgcggc
     3901 tatgtactgc gggtgaatag ctgcgctgat ggcttctaca aagaccgcta cgtgtaccgc
     3961 cacttcgcca gcgctgcact acccatcccc gaagtgttgg acatcggcga gttcagcgag
     4021 agcctgacat actgcatcag tagacgcgcc caaggcgtta ctctccaaga cctccccgaa
     4081 acagagctgc ctgctgtgtt acagcctgtc gccgaagcta tggatgctat tgccgccgcc
     4141 gacctcagtc aaaccagcgg cttcggccca ttcgggcccc aaggcatcgg ccagtacaca
     4201 acctggcggg atttcatttg cgccattgct gatccccatg tctaccactg gcagaccgtg
     4261 atggacgaca ccgtgtccgc cagcgtagct caagccctgg acgaactgat gctgtgggcc
     4321 gaagactgtc ccgaggtgcg ccacctcgtc catgccgact tcggcagcaa caacgtcctg
     4381 accgacaacg gccgcatcac cgccgtaatc gactggtccg aagctatgtt cggggacagt
     4441 cagtacgagg tggccaacat cttcttctgg cggccctggc tggcttgcat ggagcagcag
     4501 actcgctact tcgagcgccg gcatcccgag ctggccggca gccctcgtct gcgagcctac
     4561 atgctgcgca tcggcctgga tcagctctac cagagcctcg tggacggcaa cttcgacgat
     4621 gctgcctggg ctcaaggccg ctgcgatgcc atcgtccgca gcggggccgg caccgtcggt
     4681 cgcacacaaa tcgctcgccg gagcgcagcc gtatggaccg acggctgcgt cgaggtgctg
     4741 gccgacagcg gcaaccgccg gcccagtaca cgaccgcgcg ctaaggaggt aggtcgagtt
     4801 tagactctag ccatcacgat ggccgcaata aaatatcttt attttcatta catctgtgtg
     4861 ttggtttttt gtgtgaatcg atagcgataa ggatcctctt tgcgcttgcg ttttcccttg
     4921 tccagatagc ccagtagctg acattcatcc ggggtcagca ccgtttctgc ggactggctt
     4981 tctacgtaat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     5041 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     5101 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     5161 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     5221 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     5281 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     5341 ctttcaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     5401 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     5461 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     5521 ataacactgc ggccaactta cttctgacaa ctatcggagg accgaaggag ctaaccgctt
     5581 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     5641 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     5701 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     5761 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     5821 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     5881 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     5941 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaattcg
     6001 aaatgaccga ccaagcgacg cccaaccggt atcagctcac tcaaaggcgg taatacggtt
     6061 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     6121 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     6181 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     6241 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     6301 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     6361 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6421 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6481 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6541 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     6601 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6661 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6721 gcgcagaaaa aaaggatttc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6781 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6841 ctagatcctt ttatagtccg gaaatacagg aacgcacgct ggatggccct tcgctgggat
     6901 ggtgaaacca tgaaaaatgg cagcttcagt ggattaagtg ggggtaatgt ggcctgtacc
     6961 ctctggttgc ataggtattc atacggttaa aatttatcag gcgcgattgc ggcagttttt
     7021 cgggtggttt gttgccattt ttacctgtct gctgccgtga tcgcgctgaa cgcgttttag
     7081 cggtgcgtac aattaaggga ttatggtaaa tccacttact gtctgccctc gtagccatcg
     7141 agataaaccg cagtactccg gccacgatgc gtccggcgta gaggatcgag atct
//
