LOCUS       sequence-486818-        7041 bp    DNA     circular SYN 22-SEP-2025
DEFINITION  Express BRAF-NanoLuc Fusion Protein in Mammalian Cells under a CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7041)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7041)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 22, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: NanoBRET Assay Vector, BRAF-NanoLuc
FEATURES             Location/Qualifiers
     source          1..7041
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             321..833
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(921..939)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    976..1097
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(987..1006)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1041..1060
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        1342..1699
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1342..1533
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1365..1385)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1550..1685
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        1714..1761
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             1793..2587
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1847..1866)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2457..2476
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2651..2699
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(2938..3526)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3018..3037)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    3642..3925
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     enhancer        4116..4495
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4496..4699
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4650..4670
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          4835..4967
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     4950..4969
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     5011..5030
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5011..5029
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      5041..5050
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 ttatggtggg acgaggatac ctgtctccag atctcagtaa ggtacggagt aactgtccaa
       61 aagccatgaa gagattaatg gcagagtgcc tcaaaaagaa aagagatgag agaccactct
      121 ttccccaaat tctcgcctct attgagctgc tggcccgctc attgccaaaa attcaccgca
      181 gtgcatcaga accctccttg aatcgggctg gtttccaaac agaggatttt agtctatatg
      241 cttgtgcttc tccaaaaaca cccatccagg cagggggata tggtgcgttt cctgtccacg
      301 tttctctcgg ctcgagcggc gtcttcacac tcgaagattt cgttggggac tggcgacaga
      361 cagccggcta caacctggac caagtccttg aacagggagg tgtgtccagt ttgtttcaga
      421 atctcggggt gtccgtaact ccgatccaaa ggattgtcct gagcggtgaa aatgggctga
      481 agatcgacat ccatgtcatc atcccgtatg aaggtctgag cggcgaccaa atgggccaga
      541 tcgaaaaaat ttttaaggtg gtgtaccctg tggatgatca tcactttaag gtgatcctgc
      601 actatggcac actggtaatc gacggggtta cgccgaacat gatcgactat ttcggacggc
      661 cgtatgaagg catcgccgtg ttcgacggca aaaagatcac tgtaacaggg accctgtgga
      721 acggcaacaa aattatcgac gagcgcctga tcaaccccga cggctccctg ctgttccgag
      781 taaccatcaa cggagtgacc ggctggcggc tgtgcgaacg cattctggcg taaggccgcg
      841 actctagagt cgacctgcag gcatgcaagc tgatccggct gctaacaaag cccgaaagga
      901 agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg gggcggccgc
      961 ttcgagcaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     1021 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     1081 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     1141 agatgtggga ggttttttta agcaagtaaa acctctacaa atgtggtaaa atcgaatttt
     1201 aacaaaatat taacgcttac aatttcctga tgcggtattt tctccttacg catctgtgcg
     1261 gtatttcaca ccgcatacgc ggatctgcgc agcaccatgg cctgaaataa cctctgaaag
     1321 aggaacttgg ttaggtacct tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt
     1381 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     1441 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     1501 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     1561 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     1621 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     1681 aggcctaggc ttttgcaaaa agcttaatta actgttgaca attaatcatc ggcatagtat
     1741 atcggcatag tataatacga caaggtgagg aactaaaccc aggaggcaga tcatgattga
     1801 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     1861 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     1921 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga
     1981 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     2041 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     2101 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     2161 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     2221 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     2281 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga
     2341 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     2401 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     2461 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     2521 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     2581 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     2641 cgatggccgc aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga
     2701 atcgatagcg ataaggatcc tctttgcgct tgcgttttcc cttgtccaga tagcccagta
     2761 gctgacattc atccggggtc agcaccgttt ctgcggactg gctttctacc cggtatcagc
     2821 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     2881 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     2941 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3001 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3061 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3121 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3181 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3241 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     3301 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     3361 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     3421 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     3481 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tttcaagaag atcctttgat
     3541 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     3601 gagattatca aaaaggatct tcacctagat ccttttatag tccggaaata caggaacgca
     3661 cgctggatgg cccttcgctg ggatggtgaa accatgaaaa atggcagctt cagtggatta
     3721 agtgggggta atgtggcctg taccctctgg ttgcataggt attcatacgg ttaaaattta
     3781 tcaggcgcga ttgcggcagt ttttcgggtg gtttgttgcc atttttacct gtctgctgcc
     3841 gtgatcgcgc tgaacgcgtt ttagcggtgc gtacaattaa gggattatgg taaatccact
     3901 tactgtctgc cctcgtagcc atcgagataa accgcagtac tccggccacg atgcgtccgg
     3961 cgtagaggat cgagatcttc aatattggcc attagccata ttattcattg gttatatagc
     4021 ataaatcaat attggctatt ggccattgca tacgttgtat ctatatcata atatgtacat
     4081 ttatattggc tcatgtccaa tatgaccgcc atgttggcat tgattattga ctagttatta
     4141 atagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata
     4201 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat
     4261 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga
     4321 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtccgcc
     4381 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
     4441 acgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat
     4501 gcggttttgg cagtacacca atgggcgtgg atagcggttt gactcacggg gatttccaag
     4561 tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc
     4621 aaaatgtcgt aataaccccg ccccgttgac gcaaatgggc ggtaggcgtg tacggtggga
     4681 ggtctatata agcagagctg gtttagtgaa ccgtcagatc actagaagct ttattgcggt
     4741 agtttatcac agttaaattg ctaacgcagt cagtgcttct gacacaacag tctcgaactt
     4801 aagctgcaga agttggtcgt gaggcactgg gcaggtaagt atcaaggtta caagacaggt
     4861 ttaaggagac caatagaaac tgggcttgtc gagacagaga agactcttgc gtttctgata
     4921 ggcacctatt ggtcttactg acatccactt tgcctttctc tccacaggtg tccactccca
     4981 gttcaattac agctcttaag gctagagtat taatacgact cactataggg ctagcgatcg
     5041 ccatggcggc gctgagcggt ggcggtggtg gcggcgcgga gccgggccag gctctgttca
     5101 acggggacat ggagcccgag gccggcgccg gcgccggcgc cgcggcctct tcggctgcgg
     5161 accctgccat tccggaggag gtgtggaata tcaaacaaat gattaagttg acacaggaac
     5221 atatagaggc cctattggac aaatttggtg gggagcataa tccaccatca atatatctgg
     5281 aggcctatga agaatacacc agcaagctag atgcactcca acaaagagaa caacagttat
     5341 tggaatctct ggggaacgga actgattttt ctgtttctag ctctgcatca atggataccg
     5401 ttacatcttc ttcctcttct agcctttcag tgctaccttc atctctttca gtttttcaaa
     5461 atcccacaga tgtggcacgg agcaacccca agtcaccaca aaaacctatc gttagagtct
     5521 tcctgcccaa caaacagagg acagtggtac ctgcaaggtg tggagttaca gtccgagaca
     5581 gtctaaagaa agcactgatg atgagaggtc taatcccaga gtgctgtgct gtttacagaa
     5641 ttcaggatgg agagaagaaa ccaattggtt gggacactga tatttcctgg cttactggag
     5701 aagaattgca tgtggaagtg ttggagaatg ttccacttac aacacacaac tttgtacgaa
     5761 aaacgttttt caccttagca ttttgtgact tttgtcgaaa gctgcttttc cagggtttcc
     5821 gctgtcaaac atgtggttat aaatttcacc agcgttgtag tacagaagtt ccactgatgt
     5881 gtgttaatta tgaccaactt gatttgctgt ttgtctccaa gttctttgaa caccacccaa
     5941 taccacagga agaggcgtcc ttagcagaga ctgccctaac atctggatca tccccttccg
     6001 cacccgcctc ggactctatt gggccccaaa ttctcaccag tccgtctcct tcaaaatcca
     6061 ttccaattcc acagcccttc cgaccagcag atgaagatca tcgaaatcaa tttgggcaac
     6121 gagaccgatc ctcatcagct cccaatgtgc atataaacac aatagaacct gtcaatattg
     6181 atgacttgat tagagaccaa ggatttcgtg gtgatggagg atcaaccaca ggtttgtctg
     6241 ctaccccccc tgcctcatta cctggctcac taactaacgt gaaagcctta cagaaatctc
     6301 caggacctca gcgagaaagg aagtcatctt catcctcaga agacaggaat cgaatgaaaa
     6361 cacttggtag acgggactcg agtgatgatt gggagattcc tgatgggcag attacagtgg
     6421 gacaaagaat tggatctgga tcatttggaa cagtctacaa gggaaagtgg catggtgatg
     6481 tggcagtgaa aatgttgaat gtgacagcac ctacacctca gcagttacaa gccttcaaaa
     6541 atgaagtagg agtactcagg aaaacacgac atgtgaatat cctactcttc atgggctatt
     6601 ccacaaagcc acaactggct attgttaccc agtggtgtga gggctccagc ttgtatcacc
     6661 atctccatat cattgagacc aaatttgaga tgatcaaact tatagatatt gcacgacaga
     6721 ctgcacaggg catggattac ttacacgcca agtcaatcat ccacagagac ctcaagagta
     6781 ataatatatt tcttcatgaa gacctcacag taaaaatagg tgattttggt ctagctacag
     6841 tgaaatctcg atggagtggg tcccatcagt ttgaacagtt gtctggatcc attttgtgga
     6901 tggcaccaga agtcatcaga atgcaagata aaaatccata cagctttcag tcagatgtat
     6961 atgcatttgg aattgttctg tatgaattga tgactggaca gttaccttat tcaaacatca
     7021 acaacaggga ccagataatt t
//
