LOCUS       sequence-475044-        7041 bp    DNA     circular SYN 25-JUN-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)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 7041)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 25, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: NanoBRET Assay Vector, BRAF-NanoLuc
FEATURES             Location/Qualifiers
     source          1..7041
                     /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"
     regulatory      1063..1072
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3384..3896
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(3984..4002)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    4039..4160
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4050..4069)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4104..4123
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        4405..4762
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4405..4596
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4428..4448)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4613..4748
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4777..4824
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4856..5650
                     /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(4910..4929)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5520..5539
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5714..5762
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(6001..6589)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6081..6100)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6705..6988
                     /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 ggctagcgat cgccatggcg gcgctgagcg
     1081 gtggcggtgg tggcggcgcg gagccgggcc aggctctgtt caacggggac atggagcccg
     1141 aggccggcgc cggcgccggc gccgcggcct cttcggctgc ggaccctgcc attccggagg
     1201 aggtgtggaa tatcaaacaa atgattaagt tgacacagga acatatagag gccctattgg
     1261 acaaatttgg tggggagcat aatccaccat caatatatct ggaggcctat gaagaataca
     1321 ccagcaagct agatgcactc caacaaagag aacaacagtt attggaatct ctggggaacg
     1381 gaactgattt ttctgtttct agctctgcat caatggatac cgttacatct tcttcctctt
     1441 ctagcctttc agtgctacct tcatctcttt cagtttttca aaatcccaca gatgtggcac
     1501 ggagcaaccc caagtcacca caaaaaccta tcgttagagt cttcctgccc aacaaacaga
     1561 ggacagtggt acctgcaagg tgtggagtta cagtccgaga cagtctaaag aaagcactga
     1621 tgatgagagg tctaatccca gagtgctgtg ctgtttacag aattcaggat ggagagaaga
     1681 aaccaattgg ttgggacact gatatttcct ggcttactgg agaagaattg catgtggaag
     1741 tgttggagaa tgttccactt acaacacaca actttgtacg aaaaacgttt ttcaccttag
     1801 cattttgtga cttttgtcga aagctgcttt tccagggttt ccgctgtcaa acatgtggtt
     1861 ataaatttca ccagcgttgt agtacagaag ttccactgat gtgtgttaat tatgaccaac
     1921 ttgatttgct gtttgtctcc aagttctttg aacaccaccc aataccacag gaagaggcgt
     1981 ccttagcaga gactgcccta acatctggat catccccttc cgcacccgcc tcggactcta
     2041 ttgggcccca aattctcacc agtccgtctc cttcaaaatc cattccaatt ccacagccct
     2101 tccgaccagc agatgaagat catcgaaatc aatttgggca acgagaccga tcctcatcag
     2161 ctcccaatgt gcatataaac acaatagaac ctgtcaatat tgatgacttg attagagacc
     2221 aaggatttcg tggtgatgga ggatcaacca caggtttgtc tgctaccccc cctgcctcat
     2281 tacctggctc actaactaac gtgaaagcct tacagaaatc tccaggacct cagcgagaaa
     2341 ggaagtcatc ttcatcctca gaagacagga atcgaatgaa aacacttggt agacgggact
     2401 cgagtgatga ttgggagatt cctgatgggc agattacagt gggacaaaga attggatctg
     2461 gatcatttgg aacagtctac aagggaaagt ggcatggtga tgtggcagtg aaaatgttga
     2521 atgtgacagc acctacacct cagcagttac aagccttcaa aaatgaagta ggagtactca
     2581 ggaaaacacg acatgtgaat atcctactct tcatgggcta ttccacaaag ccacaactgg
     2641 ctattgttac ccagtggtgt gagggctcca gcttgtatca ccatctccat atcattgaga
     2701 ccaaatttga gatgatcaaa cttatagata ttgcacgaca gactgcacag ggcatggatt
     2761 acttacacgc caagtcaatc atccacagag acctcaagag taataatata tttcttcatg
     2821 aagacctcac agtaaaaata ggtgattttg gtctagctac agtgaaatct cgatggagtg
     2881 ggtcccatca gtttgaacag ttgtctggat ccattttgtg gatggcacca gaagtcatca
     2941 gaatgcaaga taaaaatcca tacagctttc agtcagatgt atatgcattt ggaattgttc
     3001 tgtatgaatt gatgactgga cagttacctt attcaaacat caacaacagg gaccagataa
     3061 tttttatggt gggacgagga tacctgtctc cagatctcag taaggtacgg agtaactgtc
     3121 caaaagccat gaagagatta atggcagagt gcctcaaaaa gaaaagagat gagagaccac
     3181 tctttcccca aattctcgcc tctattgagc tgctggcccg ctcattgcca aaaattcacc
     3241 gcagtgcatc agaaccctcc ttgaatcggg ctggtttcca aacagaggat tttagtctat
     3301 atgcttgtgc ttctccaaaa acacccatcc aggcaggggg atatggtgcg tttcctgtcc
     3361 acgtttctct cggctcgagc ggcgtcttca cactcgaaga tttcgttggg gactggcgac
     3421 agacagccgg ctacaacctg gaccaagtcc ttgaacaggg aggtgtgtcc agtttgtttc
     3481 agaatctcgg ggtgtccgta actccgatcc aaaggattgt cctgagcggt gaaaatgggc
     3541 tgaagatcga catccatgtc atcatcccgt atgaaggtct gagcggcgac caaatgggcc
     3601 agatcgaaaa aatttttaag gtggtgtacc ctgtggatga tcatcacttt aaggtgatcc
     3661 tgcactatgg cacactggta atcgacgggg ttacgccgaa catgatcgac tatttcggac
     3721 ggccgtatga aggcatcgcc gtgttcgacg gcaaaaagat cactgtaaca gggaccctgt
     3781 ggaacggcaa caaaattatc gacgagcgcc tgatcaaccc cgacggctcc ctgctgttcc
     3841 gagtaaccat caacggagtg accggctggc ggctgtgcga acgcattctg gcgtaaggcc
     3901 gcgactctag agtcgacctg caggcatgca agctgatccg gctgctaaca aagcccgaaa
     3961 ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcggc
     4021 cgcttcgagc agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc
     4081 agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta
     4141 taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg
     4201 gggagatgtg ggaggttttt ttaagcaagt aaaacctcta caaatgtggt aaaatcgaat
     4261 tttaacaaaa tattaacgct tacaatttcc tgatgcggta ttttctcctt acgcatctgt
     4321 gcggtatttc acaccgcata cgcggatctg cgcagcacca tggcctgaaa taacctctga
     4381 aagaggaact tggttaggta ccttctgagg cggaaagaac cagctgtgga atgtgtgtca
     4441 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4501 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     4561 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4621 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4681 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     4741 tggaggccta ggcttttgca aaaagcttaa ttaactgttg acaattaatc atcggcatag
     4801 tatatcggca tagtataata cgacaaggtg aggaactaaa cccaggaggc agatcatgat
     4861 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     4921 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     4981 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     5041 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     5101 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     5161 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     5221 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     5281 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     5341 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     5401 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     5461 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     5521 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     5581 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     5641 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     5701 tcacgatggc cgcaataaaa tatctttatt ttcattacat ctgtgtgttg gttttttgtg
     5761 tgaatcgata gcgataagga tcctctttgc gcttgcgttt tcccttgtcc agatagccca
     5821 gtagctgaca ttcatccggg gtcagcaccg tttctgcgga ctggctttct acccggtatc
     5881 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     5941 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     6001 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     6061 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     6121 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     6181 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     6241 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     6301 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     6361 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     6421 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
     6481 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     6541 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatttcaag aagatccttt
     6601 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     6661 catgagatta tcaaaaagga tcttcaccta gatcctttta tagtccggaa atacaggaac
     6721 gcacgctgga tggcccttcg ctgggatggt gaaaccatga aaaatggcag cttcagtgga
     6781 ttaagtgggg gtaatgtggc ctgtaccctc tggttgcata ggtattcata cggttaaaat
     6841 ttatcaggcg cgattgcggc agtttttcgg gtggtttgtt gccattttta cctgtctgct
     6901 gccgtgatcg cgctgaacgc gttttagcgg tgcgtacaat taagggatta tggtaaatcc
     6961 acttactgtc tgccctcgta gccatcgaga taaaccgcag tactccggcc acgatgcgtc
     7021 cggcgtagag gatcgagatc t
//
