LOCUS       sequence-425979-        6528 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of full-length human RAB7A. N-terminal NanoLuc 
            luciferase tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6528)
  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 6528)
  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_RAB7A:1-207
FEATURES             Location/Qualifiers
     source          1..6528
                     /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(2296..2314)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2346..2467
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2357..2376)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2411..2430
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        2712..3069
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2712..2903
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2735..2755)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2920..3055
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3100..4137
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4116..4124
                     /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    4161..4209
                     /note="synthetic polyadenylation signal"
     promoter        4365..4469
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4470..4538,4539..5330)
                     /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(4688..4707)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5488..6076)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5568..5587)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6192..6475
                     /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 acctctagga
     1621 agaaagtgtt gctgaaggtt atcatcctgg gagattctgg agtcgggaag acatcactca
     1681 tgaaccagta tgtgaataag aaattcagca atcagtacaa agccacaata ggagctgact
     1741 ttctgaccaa ggaggtgatg gtggatgaca ggctagtcac aatgcagata tgggacacag
     1801 caggacagga acggttccag tctctcggtg tggccttcta cagaggtgca gactgctgcg
     1861 ttctggtatt tgatgtgact gcccccaaca cattcaaaac cctagatagc tggagagatg
     1921 agtttctcat ccaggccagt ccccgagatc ctgaaaactt cccatttgtt gtgttgggaa
     1981 acaagattga cctcgaaaac agacaagtgg ccacaaagcg ggcacaggcc tggtgctaca
     2041 gcaaaaacaa cattccctac tttgagacca gtgccaagga ggccatcaac gtggagcagg
     2101 cgttccagac gattgcacgg aatgcactta agcaggaaac ggaggtggag ctgtacaacg
     2161 aatttcctga acctatcaaa ctggacaaga atgaccgggc caaggcctcg gcagaaagct
     2221 gcagttgcta ataatctaga tttgggccca attcctgcag gattttgcgg ccgcttgctg
     2281 agttggctgc tgccaccgct gagcaataac tagcataacc ccttggccgc ttcgagcaga
     2341 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
     2401 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     2461 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg agatgtggga
     2521 ggttttttta agcaagtaaa acctctacaa atgtggtaaa atcgaatttt aacaaaatat
     2581 taacgcttac aatttcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
     2641 ccgcatacgc ggatctgcgc agcaccatgg cctgaaataa cctctgaaag aggaacttgg
     2701 ttaggtacct tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga
     2761 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2821 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     2881 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     2941 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     3001 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     3061 ttttgcaaaa agctcgattc ttctgacact agcgccacca tgaagaagcc cgaactcacc
     3121 gctaccagcg ttgaaaaatt tctcatcgag aagttcgaca gtgtgagcga cctgatgcag
     3181 ttgtcggagg gcgaagagag ccgagccttc agcttcgatg tcggcggacg cggctatgta
     3241 ctgcgggtga atagctgcgc tgatggcttc tacaaagacc gctacgtgta ccgccacttc
     3301 gccagcgctg cactacccat ccccgaagtg ttggacatcg gcgagttcag cgagagcctg
     3361 acatactgca tcagtagacg cgcccaaggc gttactctcc aagacctccc cgaaacagag
     3421 ctgcctgctg tgttacagcc tgtcgccgaa gctatggatg ctattgccgc cgccgacctc
     3481 agtcaaacca gcggcttcgg cccattcggg ccccaaggca tcggccagta cacaacctgg
     3541 cgggatttca tttgcgccat tgctgatccc catgtctacc actggcagac cgtgatggac
     3601 gacaccgtgt ccgccagcgt agctcaagcc ctggacgaac tgatgctgtg ggccgaagac
     3661 tgtcccgagg tgcgccacct cgtccatgcc gacttcggca gcaacaacgt cctgaccgac
     3721 aacggccgca tcaccgccgt aatcgactgg tccgaagcta tgttcgggga cagtcagtac
     3781 gaggtggcca acatcttctt ctggcggccc tggctggctt gcatggagca gcagactcgc
     3841 tacttcgagc gccggcatcc cgagctggcc ggcagccctc gtctgcgagc ctacatgctg
     3901 cgcatcggcc tggatcagct ctaccagagc ctcgtggacg gcaacttcga cgatgctgcc
     3961 tgggctcaag gccgctgcga tgccatcgtc cgcagcgggg ccggcaccgt cggtcgcaca
     4021 caaatcgctc gccggagcgc agccgtatgg accgacggct gcgtcgaggt gctggccgac
     4081 agcggcaacc gccggcccag tacacgaccg cgcgctaagg aggtaggtcg agtttagact
     4141 ctagccatca cgatggccgc aataaaatat ctttattttc attacatctg tgtgttggtt
     4201 ttttgtgtga atcgatagcg ataaggatcc tctttgcgct tgcgttttcc cttgtccaga
     4261 tagcccagta gctgacattc atccggggtc agcaccgttt ctgcggactg gctttctacg
     4321 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     4381 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     4441 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     4501 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     4561 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     4621 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttca
     4681 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     4741 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     4801 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     4861 ctgcggccaa cttacttctg acaactatcg gaggaccgaa ggagctaacc gcttttttgc
     4921 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     4981 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     5041 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     5101 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     5161 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     5221 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     5281 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ttcgaaatga
     5341 ccgaccaagc gacgcccaac cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5401 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5461 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5521 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5581 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5641 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5701 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5761 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5821 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5881 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     5941 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     6001 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     6061 aaaaaaagga tttcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     6121 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     6181 ccttttatag tccggaaata caggaacgca cgctggatgg cccttcgctg ggatggtgaa
     6241 accatgaaaa atggcagctt cagtggatta agtgggggta atgtggcctg taccctctgg
     6301 ttgcataggt attcatacgg ttaaaattta tcaggcgcga ttgcggcagt ttttcgggtg
     6361 gtttgttgcc atttttacct gtctgctgcc gtgatcgcgc tgaacgcgtt ttagcggtgc
     6421 gtacaattaa gggattatgg taaatccact tactgtctgc cctcgtagcc atcgagataa
     6481 accgcagtac tccggccacg atgcgtccgg cgtagaggat cgagatct
//
