LOCUS       sequence-257754-        7874 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of Flag tagged BRAF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7874)
  AUTHORS   Noeparast A, Teugels E, Giron P, Verschelden G, De Brakeleer S, 
            Decoster L, De Greve J
  TITLE     Non-V600 BRAF mutations recurrently found in lung cancer predict 
            sensitivity to the combination of Trametinib and Dabrafenib.
  JOURNAL   Oncotarget. 2016 Aug 26;8(36):60094-60108. doi: 
            10.18632/oncotarget.11635. eCollection 2017 Sep 1.
  PUBMED    28947956
REFERENCE   2  (bases 1 to 7874)
  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: 
            "10.18632/oncotarget.11635"; journalName: "Oncotarget"; date: 
            "2016-08-26- 26"; volume: "8"; issue: "36"; pages: "60094-60108"
COMMENT     Sequence Label: p3xFLAG CMV14 / BRAF-V600K
FEATURES             Location/Qualifiers
     source          1..7874
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             2937..3002
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    3013..3635
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3212..3231)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(3659..3679)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3677..3974
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             4028..4822
                     /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(4082..4101)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4692..4711
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(5351..5939)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5431..5450)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6110..6901,6902..6970))
                     /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     6733..6752
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6971..7075)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      7102..7557
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7189..7208)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7399..7420
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7683..7705
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7697..7714
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7698..7714
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agaattaagc ttgcggccgc atggcggcgc tgagcggtgg cggtggtggc ggcgcggagc
      661 cgggccaggc tctgttcaac ggggacatgg agcccgaggc cggcgccggc gccggcgccg
      721 cggcctcttc ggctgcggac cctgccattc cggaggaggt gtggaatatc aaacaaatga
      781 ttaagttgac acaggaacat atagaggccc tattggacaa atttggtggg gagcataatc
      841 caccatcaat atatctggag gcctatgaag aatacaccag caagctagat gcactccaac
      901 aaagagaaca acagttattg gaatctctgg ggaacggaac tgatttttct gtttctagct
      961 ctgcatcaat ggataccgtt acatcttctt cctcttctag cctttcagtg ctaccttcat
     1021 ctctttcagt ttttcaaaat cccacagatg tggcacggag caaccccaag tcaccacaaa
     1081 aacctatcgt tagagtcttc ctgcccaaca aacagaggac agtggtacct gcaaggtgtg
     1141 gagttacagt ccgagacagt ctaaagaaag cactgatgat gagaggtcta atcccagagt
     1201 gctgtgctgt ttacagaatt caggatggag agaagaaacc aattggttgg gacactgata
     1261 tttcctggct tactggagaa gaattgcatg tggaagtgtt ggagaatgtt ccacttacaa
     1321 cacacaactt tgtacgaaaa acgtttttca ccttagcatt ttgtgacttt tgtcgaaagc
     1381 tgcttttcca gggtttccgc tgtcaaacat gtggttataa atttcaccag cgttgtagta
     1441 cagaagttcc actgatgtgt gttaattatg accaacttga tttgctgttt gtctccaagt
     1501 tctttgaaca ccacccaata ccacaggaag aggcgtcctt agcagagact gccctaacat
     1561 ctggatcatc cccttccgca cccgcctcgg actctattgg gccccaaatt ctcaccagtc
     1621 cgtctccttc aaaatccatt ccaattccac agcccttccg accagcagat gaagatcatc
     1681 gaaatcaatt tgggcaacga gaccgatcct catcagctcc caatgtgcat ataaacacaa
     1741 tagaacctgt caatattgat gacttgatta gagaccaagg atttcgtggt gatggaggat
     1801 caaccacagg tttgtctgct accccccctg cctcattacc tggctcacta actaacgtga
     1861 aagccttaca gaaatctcca ggacctcagc gagaaaggaa gtcatcttca tcctcagaag
     1921 acaggaatcg aatgaaaaca cttggtagac gggactcgag tgatgattgg gagattcctg
     1981 atgggcagat tacagtggga caaagaattg gatctggatc atttggaaca gtctacaagg
     2041 gaaagtggca tggtgatgtg gcagtgaaaa tgttgaatgt gacagcacct acacctcagc
     2101 agttacaagc cttcaaaaat gaagtaggag tactcaggaa aacacgacat gtgaatatcc
     2161 tactcttcat gggctattcc acaaagccac aactggctat tgttacccag tggtgtgagg
     2221 gctccagctt gtatcaccat ctccatatca ttgagaccaa atttgagatg atcaaactta
     2281 tagatattgc acgacagact gcacagggca tggattactt acacgccaag tcaatcatcc
     2341 acagagacct caagagtaat aatatatttc ttcatgaaga cctcacagta aaaataggtg
     2401 attttggtct agctacaaag aaatctcgat ggagtgggtc ccatcagttt gaacagttgt
     2461 ctggatccat tttgtggatg gcaccagaag tcatcagaat gcaagataaa aatccataca
     2521 gctttcagtc agatgtatat gcatttggaa ttgttctgta tgaattgatg actggacagt
     2581 taccttattc aaacatcaac aacagggacc agataatttt tatggtggga cgaggatacc
     2641 tgtctccaga tctcagtaag gtacggagta actgtccaaa agccatgaag agattaatgg
     2701 cagagtgcct caaaaagaaa agagatgaga gaccactctt tccccaaatt ctcgcctcta
     2761 ttgagctgct ggcccgctca ttgccaaaaa ttcaccgcag tgcatcagaa ccctccttga
     2821 atcgggctgg tttccaaaca gaggatttta gtctatatgc ttgtgcttct ccaaaaacac
     2881 ccatccaggc agggggatat ggtgcgtttc ctgtccactc tagaggatcc cgggctgact
     2941 acaaagacca tgacggtgat tataaagatc atgacatcga ctacaaggat gacgatgaca
     3001 agtagtgatc ccgggtggca tccctgtgac ccctccccag tgcctctcct ggccctggaa
     3061 gttgccactc cagtgcccac cagccttgtc ctaataaaat taagttgcat cattttgtct
     3121 gactaggtgt ccttctataa tattatgggg tggagggggg tggtatggag caaggggcaa
     3181 gttgggaaga caacctgtag ggcctgcggg gtctattggg aaccaagctg gagtgcagtg
     3241 gcacaatctt ggctcactgc aatctccgcc tcctgggttc aagcgattct cctgcctcag
     3301 cctcccgagt tgttgggatt ccaggcatgc atgaccaggc tcagctaatt tttgtttttt
     3361 tggtagagac ggggtttcac catattggcc aggctggtct ccaactccta atctcaggtg
     3421 atctacccac cttggcctcc caaattgctg ggattacagg cgtgaaccac tgctcccttc
     3481 cctgtccttc tgattttaaa ataactatac cagcaggagg acgtccagac acagcatagg
     3541 ctacctggcc atgcccaacc ggtgggacat ttgagttgct tgcttggcac tgtcctctca
     3601 tgcgttgggt ccactcagta gatgcctgtt gaattgggta cgcggccagc ttggctgtgg
     3661 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     3721 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     3781 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3841 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3901 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     3961 ggaggctttt ttggaggaat tgatcagctt gggatctgat caagagacag gatgaggatc
     4021 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     4081 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     4141 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     4201 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     4261 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     4321 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     4381 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     4441 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     4501 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat
     4561 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     4621 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     4681 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     4741 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     4801 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     4861 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     4921 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     4981 ttcttcgccc accccgggct cgatcccctc gcgagttggt tcagctgctg cctgaggctg
     5041 gacgacctcg cggagttcta ccggcagtgc aaatccgtcg gcatccagga aaccagcagc
     5101 ggctatccgc gcatccatgc ccccgaactg caggagtggg gaggcacgat ggccgctttg
     5161 gtcgacccgg acgggacgct cctgcgcctg atacagaacg aattgcttgc aggcatctca
     5221 tgagtgtgtc ttcccgtttt ccgcctgagg tcactgcgtg gatggagcgc tggcgcctgc
     5281 tgcgcgacgg cgagctgctc accacccact cgccaagctg gaaccgtaaa aaggccgcgt
     5341 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5401 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5461 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5521 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5581 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     5641 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     5701 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5761 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     5821 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5881 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5941 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6001 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6061 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6121 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6181 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6241 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     6301 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6361 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     6421 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     6481 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     6541 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     6601 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     6661 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     6721 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     6781 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6841 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     6901 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     6961 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     7021 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     7081 ttccccgaaa agtgccacct gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     7141 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     7201 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     7261 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     7321 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     7381 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     7441 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     7501 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttcca
     7561 ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt
     7621 acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt
     7681 ttcccagtca cgacgttgta aaacgacggc cagtgccaag ctgatctata cattgaatca
     7741 atattggcaa ttagccatat tagtcattgg ttatatagca taaatcaata ttggctattg
     7801 gccattgcat acgttgtatc tatatcataa tatgtacatt tatattggct catgtccaat
     7861 atgaccgcca tgtt
//
