LOCUS       sequence-423316-        6849 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human MAGEA3. N-terminal NanoLuc
            luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6849)
  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 6849)
  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-N_MAGEA3:1-314
FEATURES             Location/Qualifiers
     source          1..6849
                     /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             1065..1577
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     primer_bind     complement(2617..2635)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2667..2788
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2678..2697)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2732..2751
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3033..3390
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3033..3224
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3056..3076)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3241..3376
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3421..4458
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4437..4445
                     /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    4482..4530
                     /note="synthetic polyadenylation signal"
     promoter        4686..4790
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4791..4859,4860..5651)
                     /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(5009..5028)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5809..6397)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5889..5908)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6513..6796
                     /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 ggctagcgct caccatggtc ttcacactcg
     1081 aagatttcgt tggggactgg cgacagacag ccggctacaa cctggaccaa gtccttgaac
     1141 agggaggtgt gtccagtttg tttcagaatc tcggggtgtc cgtaactccg atccaaagga
     1201 ttgtcctgag cggtgaaaat gggctgaaga tcgacatcca tgtcatcatc ccgtatgaag
     1261 gtctgagcgg cgaccaaatg ggccagatcg aaaaaatttt taaggtggtg taccctgtgg
     1321 atgatcatca ctttaaggtg atcctgcact atggcacact ggtaatcgac ggggttacgc
     1381 cgaacatgat cgactatttc ggacggccgt atgaaggcat cgccgtgttc gacggcaaaa
     1441 agatcactgt aacagggacc ctgtggaacg gcaacaaaat tatcgacgag cgcctgatca
     1501 accccgacgg ctccctgctg ttccgagtaa ccatcaacgg agtgaccggc tggcggctgt
     1561 gcgaacgcat tctggcgggc tcgagcggcg cgatcgcttc cgaattcatg cctcttgagc
     1621 agaggagtca gcactgcaag cctgaagaag gccttgaggc ccgaggagag gccctgggcc
     1681 tggtgggtgc gcaggctcct gctactgagg agcaggaggc tgcctcctcc tcttctactc
     1741 tagttgaagt caccctgggg gaggtgcctg ctgccgagtc accagatcct ccccagagtc
     1801 ctcagggagc ctccagcctc cccactacca tgaactaccc tctctggagc caatcctatg
     1861 aggactccag caaccaagaa gaggaggggc caagcacctt ccctgacctg gagtccgagt
     1921 tccaagcagc actcagtagg aaggtggccg agttggttca ttttctgctc ctcaagtatc
     1981 gagccaggga gccggtcaca aaggcagaaa tgctggggag tgtcgtcgga aattggcagt
     2041 atttctttcc tgtgatcttc agcaaagctt ccagttcctt gcagctggtc tttggcatcg
     2101 agctgatgga agtggacccc atcggccact tgtacatctt tgccacctgc ctgggcctct
     2161 cctacgatgg cctgctgggt gacaatcaga tcatgcccaa ggcaggcctc ctgataatcg
     2221 tcctggccat aatcgcaaga gagggcgact gtgcccctga ggagaaaatc tgggaggagc
     2281 tgagtgtgtt agaggtgttt gaggggaggg aagacagtat cttgggggat cccaagaagc
     2341 tgctcaccca acatttcgtg caggaaaact acctggagta ccggcaggtc cccggcagtg
     2401 atcctgcatg ttatgaattc ctgtggggtc caagggccct cgttgaaacc agctatgtga
     2461 aagtcctgca ccatatggta aagatcagtg gaggacctca catttcctac ccacccctgc
     2521 atgagtgggt tttgagagag ggggaagagt aataatctag atttgggccc aattcctgca
     2581 ggattttgcg gccgcttgct gagttggctg ctgccaccgc tgagcaataa ctagcataac
     2641 cccttggccg cttcgagcag acatgataag atacattgat gagtttggac aaaccacaac
     2701 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     2761 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     2821 ggttcagggg gagatgtggg aggttttttt aagcaagtaa aacctctaca aatgtggtaa
     2881 aatcgaattt taacaaaata ttaacgctta caatttcctg atgcggtatt ttctccttac
     2941 gcatctgtgc ggtatttcac accgcatacg cggatctgcg cagcaccatg gcctgaaata
     3001 acctctgaaa gaggaacttg gttaggtacc ttctgaggcg gaaagaacca gctgtggaat
     3061 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3121 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     3181 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     3241 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     3301 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     3361 ggcttttttg gaggcctagg cttttgcaaa aagctcgatt cttctgacac tagcgccacc
     3421 atgaagaagc ccgaactcac cgctaccagc gttgaaaaat ttctcatcga gaagttcgac
     3481 agtgtgagcg acctgatgca gttgtcggag ggcgaagaga gccgagcctt cagcttcgat
     3541 gtcggcggac gcggctatgt actgcgggtg aatagctgcg ctgatggctt ctacaaagac
     3601 cgctacgtgt accgccactt cgccagcgct gcactaccca tccccgaagt gttggacatc
     3661 ggcgagttca gcgagagcct gacatactgc atcagtagac gcgcccaagg cgttactctc
     3721 caagacctcc ccgaaacaga gctgcctgct gtgttacagc ctgtcgccga agctatggat
     3781 gctattgccg ccgccgacct cagtcaaacc agcggcttcg gcccattcgg gccccaaggc
     3841 atcggccagt acacaacctg gcgggatttc atttgcgcca ttgctgatcc ccatgtctac
     3901 cactggcaga ccgtgatgga cgacaccgtg tccgccagcg tagctcaagc cctggacgaa
     3961 ctgatgctgt gggccgaaga ctgtcccgag gtgcgccacc tcgtccatgc cgacttcggc
     4021 agcaacaacg tcctgaccga caacggccgc atcaccgccg taatcgactg gtccgaagct
     4081 atgttcgggg acagtcagta cgaggtggcc aacatcttct tctggcggcc ctggctggct
     4141 tgcatggagc agcagactcg ctacttcgag cgccggcatc ccgagctggc cggcagccct
     4201 cgtctgcgag cctacatgct gcgcatcggc ctggatcagc tctaccagag cctcgtggac
     4261 ggcaacttcg acgatgctgc ctgggctcaa ggccgctgcg atgccatcgt ccgcagcggg
     4321 gccggcaccg tcggtcgcac acaaatcgct cgccggagcg cagccgtatg gaccgacggc
     4381 tgcgtcgagg tgctggccga cagcggcaac cgccggccca gtacacgacc gcgcgctaag
     4441 gaggtaggtc gagtttagac tctagccatc acgatggccg caataaaata tctttatttt
     4501 cattacatct gtgtgttggt tttttgtgtg aatcgatagc gataaggatc ctctttgcgc
     4561 ttgcgttttc ccttgtccag atagcccagt agctgacatt catccggggt cagcaccgtt
     4621 tctgcggact ggctttctac gtaatggttt cttagacgtc aggtggcact tttcggggaa
     4681 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     4741 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     4801 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     4861 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     4921 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     4981 ttccaatgat gagcactttc aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     5041 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     5101 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     5161 ccataaccat gagtgataac actgcggcca acttacttct gacaactatc ggaggaccga
     5221 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     5281 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     5341 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     5401 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     5461 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     5521 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     5581 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     5641 agcattggta attcgaaatg accgaccaag cgacgcccaa ccggtatcag ctcactcaaa
     5701 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5761 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5821 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5881 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5941 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     6001 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     6061 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     6121 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     6181 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     6241 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     6301 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     6361 caagcagcag attacgcgca gaaaaaaagg atttcaagaa gatcctttga tcttttctac
     6421 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     6481 aaaaaggatc ttcacctaga tccttttata gtccggaaat acaggaacgc acgctggatg
     6541 gcccttcgct gggatggtga aaccatgaaa aatggcagct tcagtggatt aagtgggggt
     6601 aatgtggcct gtaccctctg gttgcatagg tattcatacg gttaaaattt atcaggcgcg
     6661 attgcggcag tttttcgggt ggtttgttgc catttttacc tgtctgctgc cgtgatcgcg
     6721 ctgaacgcgt tttagcggtg cgtacaatta agggattatg gtaaatccac ttactgtctg
     6781 ccctcgtagc catcgagata aaccgcagta ctccggccac gatgcgtccg gcgtagagga
     6841 tcgagatct
//
