LOCUS       sequence-425969-        7539 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human PRKCSH. C-terminal NanoLuc
            luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7539)
  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 7539)
  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_PRKCSH:1-527
FEATURES             Location/Qualifiers
     source          1..7539
                     /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             2714..3226
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(3302..3320)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3357..3478
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3368..3387)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3422..3441
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3723..4080
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3723..3914
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3746..3766)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3931..4066
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4111..5148
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             5127..5135
                     /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    5172..5220
                     /note="synthetic polyadenylation signal"
     promoter        5376..5480
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5481..5549,5550..6341)
                     /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(5699..5718)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6499..7087)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6579..6598)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    7203..7486
                     /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 caccatgctg ttgccgctgc tgctgctgct acccatgtgc tgggccgtgg aggtcaagag
     1141 gccccggggc gtctccctca ccaatcatca cttctacgat gagtccaagc ctttcacctg
     1201 cctggacggt tcggccacca tcccatttga tcaggtcaac gatgactatt gcgactgcaa
     1261 agatggctct gacgagccag gcacggctgc ctgtcctaat ggcagcttcc actgcaccaa
     1321 cactggctat aagcccctgt atatcccctc caaccgggtc aacgatggtg tttgtgactg
     1381 ctgcgatgga acagacgagt acaacagcgg cgtcatctgt gagaacacct gcaaagagaa
     1441 gggccgtaag gagagagagt ccctgcagca gatggccgag gtcacccgcg aagggttccg
     1501 tctgaagaag atccttattg aggactggaa gaaggcacgg gaggagaagc agaaaaagct
     1561 cattgagcta caggctggga agaagtctct ggaagaccag gtggagatgc tgcggacagt
     1621 gaaggaggaa gctgagaagc cagagagaga ggccaaagag cagcaccaga agctgtggga
     1681 agagcagctg gctgctgcca aggcccaaca ggagcaggag ctggcggctg atgccttcaa
     1741 ggagctggat gatgacatgg acgggacggt ctcggtgact gagctgcaga ctcacccgga
     1801 gctggacaca gatggggatg gggcgttgtc agaagcggaa gctcaggccc tcctcagtgg
     1861 ggacacacag acagacgcca cctctttcta cgaccgcgtc tgggccgcca tcagggacaa
     1921 gtaccggtcc gaggcactgc ccaccgacct tccaacacct tctgcccctg acttgacgga
     1981 gcccaaggag gagcagccgc cagtgccctc gtcgcccaca gaggaggagg aggaggagga
     2041 ggaggaggaa gaagaggctg aagaagagga ggaggaggag gattccgagg aggccccacc
     2101 gccactgtca cccccgcagc cggccagccc tgctgaggaa gacaaaatgc cgccctacga
     2161 cgagcagacg caggccttca tcgatgctgc ccaggaggcc cgcaacaagt tcgaggaggc
     2221 cgagcggtcg ctgaaggaca tggaggagtc catcaggaac ctggagcaag agatttcttt
     2281 tgactttggc cccaacgggg agtttgctta cctgtacagc cagtgctacg agctcaccac
     2341 caacgaatac gtctaccgcc tctgcccctt caagcttgtc tcgcagaaac ccaaactcgg
     2401 gggctctccc accagccttg gcacctgggg ctcatggatt ggccccgacc acgacaagtt
     2461 cagtgccatg aagtatgagc aaggcacggg ctgctggcag ggccccaacc gctccaccac
     2521 cgtgcgcctc ctgtgcggga aagagaccat ggtgaccagc accacagagc ccagtcgctg
     2581 cgagtacctc atggagctga tgacgccagc cgcctgcccg gagccaccgc ctgaagcacc
     2641 caccgaagac gaccatgacg agctctctag atttgggccc aattcctgca ggcgagctct
     2701 cggctcgagc ggcgtcttca cactcgaaga tttcgttggg gactggcgac agacagccgg
     2761 ctacaacctg gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg
     2821 ggtgtccgta actccgatcc aaaggattgt cctgagcggt gaaaatgggc tgaagatcga
     2881 catccatgtc atcatcccgt atgaaggtct gagcggcgac caaatgggcc agatcgaaaa
     2941 aatttttaag gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc tgcactatgg
     3001 cacactggta atcgacgggg ttacgccgaa catgatcgac tatttcggac ggccgtatga
     3061 aggcatcgcc gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa
     3121 caaaattatc gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat
     3181 caacggagtg accggctggc ggctgtgcga acgcattctg gcgtaaggcc gcgactctag
     3241 ggcatgcaag ctgatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
     3301 accgctgagc aataactagc ataacccctt ggggcggccg cttcgagcag acatgataag
     3361 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     3421 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     3481 caacaacaat tgcattcatt ttatgtttca ggttcagggg gagatgtggg aggttttttt
     3541 aagcaagtaa aacctctaca aatgtggtaa aatcgaattt taacaaaata ttaacgctta
     3601 caatttcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
     3661 cggatctgcg cagcaccatg gcctgaaata acctctgaaa gaggaacttg gttaggtacc
     3721 ttctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     3781 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     3841 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3901 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     3961 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
     4021 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     4081 aagctcgatt cttctgacac tagcgccacc atgaagaagc ccgaactcac cgctaccagc
     4141 gttgaaaaat ttctcatcga gaagttcgac agtgtgagcg acctgatgca gttgtcggag
     4201 ggcgaagaga gccgagcctt cagcttcgat gtcggcggac gcggctatgt actgcgggtg
     4261 aatagctgcg ctgatggctt ctacaaagac cgctacgtgt accgccactt cgccagcgct
     4321 gcactaccca tccccgaagt gttggacatc ggcgagttca gcgagagcct gacatactgc
     4381 atcagtagac gcgcccaagg cgttactctc caagacctcc ccgaaacaga gctgcctgct
     4441 gtgttacagc ctgtcgccga agctatggat gctattgccg ccgccgacct cagtcaaacc
     4501 agcggcttcg gcccattcgg gccccaaggc atcggccagt acacaacctg gcgggatttc
     4561 atttgcgcca ttgctgatcc ccatgtctac cactggcaga ccgtgatgga cgacaccgtg
     4621 tccgccagcg tagctcaagc cctggacgaa ctgatgctgt gggccgaaga ctgtcccgag
     4681 gtgcgccacc tcgtccatgc cgacttcggc agcaacaacg tcctgaccga caacggccgc
     4741 atcaccgccg taatcgactg gtccgaagct atgttcgggg acagtcagta cgaggtggcc
     4801 aacatcttct tctggcggcc ctggctggct tgcatggagc agcagactcg ctacttcgag
     4861 cgccggcatc ccgagctggc cggcagccct cgtctgcgag cctacatgct gcgcatcggc
     4921 ctggatcagc tctaccagag cctcgtggac ggcaacttcg acgatgctgc ctgggctcaa
     4981 ggccgctgcg atgccatcgt ccgcagcggg gccggcaccg tcggtcgcac acaaatcgct
     5041 cgccggagcg cagccgtatg gaccgacggc tgcgtcgagg tgctggccga cagcggcaac
     5101 cgccggccca gtacacgacc gcgcgctaag gaggtaggtc gagtttagac tctagccatc
     5161 acgatggccg caataaaata tctttatttt cattacatct gtgtgttggt tttttgtgtg
     5221 aatcgatagc gataaggatc ctctttgcgc ttgcgttttc ccttgtccag atagcccagt
     5281 agctgacatt catccggggt cagcaccgtt tctgcggact ggctttctac gtaatggttt
     5341 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     5401 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     5461 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     5521 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     5581 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     5641 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttc aaagttctgc
     5701 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     5761 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     5821 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     5881 acttacttct gacaactatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     5941 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     6001 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     6061 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     6121 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     6181 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     6241 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     6301 agatcgctga gataggtgcc tcactgatta agcattggta attcgaaatg accgaccaag
     6361 cgacgcccaa ccggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6421 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6481 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     6541 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6601 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6661 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     6721 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6781 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     6841 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     6901 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     6961 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7021 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     7081 atttcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     7141 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttata
     7201 gtccggaaat acaggaacgc acgctggatg gcccttcgct gggatggtga aaccatgaaa
     7261 aatggcagct tcagtggatt aagtgggggt aatgtggcct gtaccctctg gttgcatagg
     7321 tattcatacg gttaaaattt atcaggcgcg attgcggcag tttttcgggt ggtttgttgc
     7381 catttttacc tgtctgctgc cgtgatcgcg ctgaacgcgt tttagcggtg cgtacaatta
     7441 agggattatg gtaaatccac ttactgtctg ccctcgtagc catcgagata aaccgcagta
     7501 ctccggccac gatgcgtccg gcgtagagga tcgagatct
//
