LOCUS       sequence-423299-        6213 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human DDA1. N-terminal NanoLuc 
            luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6213)
  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 6213)
  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_DDA1:1-102
FEATURES             Location/Qualifiers
     source          1..6213
                     /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(1981..1999)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2031..2152
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2042..2061)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2096..2115
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        2397..2754
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2397..2588
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2420..2440)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2605..2740
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2785..3822
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             3801..3809
                     /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    3846..3894
                     /note="synthetic polyadenylation signal"
     promoter        4050..4154
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4155..4223,4224..5015)
                     /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(4373..4392)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5173..5761)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5253..5272)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    5877..6160
                     /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 gcagattttt
     1621 tgaaaggact gcctgtctac aacaaaagca attttagtcg atttcacgcg gactccgtgt
     1681 gcaaagcctc gaaccgacgg ccctcagtct acctgcctac ccgcgagtac ccgtctgaac
     1741 agatcatcgt gacagaaaag acaaacatcc tcctgcgcta cctgcatcag caatgggaca
     1801 aaaagaacgc tgccaagaag agagaccagg agcaagtgga gctggaaggc gagagctccg
     1861 cacctccccg caaggtggcg cggaccgaca gcccagacat gcacgaggac acttaataat
     1921 ctagatttgg gcccaattcc tgcaggattt tgcggccgct tgctgagttg gctgctgcca
     1981 ccgctgagca ataactagca taaccccttg gccgcttcga gcagacatga taagatacat
     2041 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     2101 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa
     2161 caattgcatt cattttatgt ttcaggttca gggggagatg tgggaggttt ttttaagcaa
     2221 gtaaaacctc tacaaatgtg gtaaaatcga attttaacaa aatattaacg cttacaattt
     2281 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgcggatc
     2341 tgcgcagcac catggcctga aataacctct gaaagaggaa cttggttagg taccttctga
     2401 ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     2461 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     2521 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     2581 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     2641 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     2701 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     2761 gattcttctg acactagcgc caccatgaag aagcccgaac tcaccgctac cagcgttgaa
     2821 aaatttctca tcgagaagtt cgacagtgtg agcgacctga tgcagttgtc ggagggcgaa
     2881 gagagccgag ccttcagctt cgatgtcggc ggacgcggct atgtactgcg ggtgaatagc
     2941 tgcgctgatg gcttctacaa agaccgctac gtgtaccgcc acttcgccag cgctgcacta
     3001 cccatccccg aagtgttgga catcggcgag ttcagcgaga gcctgacata ctgcatcagt
     3061 agacgcgccc aaggcgttac tctccaagac ctccccgaaa cagagctgcc tgctgtgtta
     3121 cagcctgtcg ccgaagctat ggatgctatt gccgccgccg acctcagtca aaccagcggc
     3181 ttcggcccat tcgggcccca aggcatcggc cagtacacaa cctggcggga tttcatttgc
     3241 gccattgctg atccccatgt ctaccactgg cagaccgtga tggacgacac cgtgtccgcc
     3301 agcgtagctc aagccctgga cgaactgatg ctgtgggccg aagactgtcc cgaggtgcgc
     3361 cacctcgtcc atgccgactt cggcagcaac aacgtcctga ccgacaacgg ccgcatcacc
     3421 gccgtaatcg actggtccga agctatgttc ggggacagtc agtacgaggt ggccaacatc
     3481 ttcttctggc ggccctggct ggcttgcatg gagcagcaga ctcgctactt cgagcgccgg
     3541 catcccgagc tggccggcag ccctcgtctg cgagcctaca tgctgcgcat cggcctggat
     3601 cagctctacc agagcctcgt ggacggcaac ttcgacgatg ctgcctgggc tcaaggccgc
     3661 tgcgatgcca tcgtccgcag cggggccggc accgtcggtc gcacacaaat cgctcgccgg
     3721 agcgcagccg tatggaccga cggctgcgtc gaggtgctgg ccgacagcgg caaccgccgg
     3781 cccagtacac gaccgcgcgc taaggaggta ggtcgagttt agactctagc catcacgatg
     3841 gccgcaataa aatatcttta ttttcattac atctgtgtgt tggttttttg tgtgaatcga
     3901 tagcgataag gatcctcttt gcgcttgcgt tttcccttgt ccagatagcc cagtagctga
     3961 cattcatccg gggtcagcac cgtttctgcg gactggcttt ctacgtaatg gtttcttaga
     4021 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     4081 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     4141 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     4201 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     4261 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     4321 agagttttcg ccccgaagaa cgttttccaa tgatgagcac tttcaaagtt ctgctatgtg
     4381 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     4441 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     4501 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     4561 ttctgacaac tatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     4621 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     4681 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     4741 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     4801 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     4861 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     4921 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     4981 ctgagatagg tgcctcactg attaagcatt ggtaattcga aatgaccgac caagcgacgc
     5041 ccaaccggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     5101 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     5161 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     5221 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5281 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5341 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5401 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5461 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5521 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5581 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     5641 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5701 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatttca
     5761 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5821 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt tatagtccgg
     5881 aaatacagga acgcacgctg gatggccctt cgctgggatg gtgaaaccat gaaaaatggc
     5941 agcttcagtg gattaagtgg gggtaatgtg gcctgtaccc tctggttgca taggtattca
     6001 tacggttaaa atttatcagg cgcgattgcg gcagtttttc gggtggtttg ttgccatttt
     6061 tacctgtctg ctgccgtgat cgcgctgaac gcgttttagc ggtgcgtaca attaagggat
     6121 tatggtaaat ccacttactg tctgccctcg tagccatcga gataaaccgc agtactccgg
     6181 ccacgatgcg tccggcgtag aggatcgaga tct
//
