LOCUS       sequence-372943-        7248 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Flag tagged human oncogenic BRAFV600E from bleomycin 
            resistance retroviral vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7248)
  AUTHORS   Brady DC, Crowe MS, Turski ML, Hobbs GA, Yao X, Chaikuad A, Knapp S,
            Xiao K, Campbell SL, Thiele DJ, Counter CM
  TITLE     Copper is required for oncogenic BRAF signalling and tumorigenesis.
  JOURNAL   Nature. 2014 Apr 9. doi: 10.1038/nature13180.
  PUBMED    24717435
REFERENCE   2  (bases 1 to 7248)
  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"; journalName: "Nature. 
            2014 Apr 9. doi: 10.1038/nature13180."
COMMENT     Sequence Label: pBABEbleo-Flag-BRAFV600E
FEATURES             Location/Qualifiers
     source          1..7248
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(89..109)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        94..423
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      274..409
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             483..857
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     LTR             1075..1544
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(1674..1691)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1845..2433)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1925..1944)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2593..3384,3385..3453))
                     /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     3216..3235
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3454..3558)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             3589..4058
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    4123..4322
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             4523..4939
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     4869..4891
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     4907..4923
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      4995..5004
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5004..5027
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
ORIGIN
        1 attttagtct atatgcttgt gcttctccaa aaacacccat ccaggcaggg ggatatggtg
       61 cgtttcctgt ccactgagtc gaccctgtgg aatgtgtgtc agttagggtg tggaaagtcc
      121 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
      181 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
      241 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
      301 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
      361 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
      421 aaaaagcttg ccagctgggg cgccctctgg taaggttggg aagccctgca gatctgatca
      481 ccatggccaa gttgaccagt gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg
      541 tcgagttctg gaccgaccgg ctcgggttct cccgggactt cgtggaggac gacttcgccg
      601 gtgtggtccg ggacgacgtg accctgttca tcagcgcggt ccaggaccag gtggtgccgg
      661 acaacaccct ggcctgggtg tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg
      721 aggtcgtgtc cacgaacttc cgggacgcct ccgggccggc catgaccgag atcggcgagc
      781 agccgtgggg gcgggagttc gccctgcgcg acccggccgg caactgcgtg cacttcgtgg
      841 ccgaggagca ggactgaccg acgccgacca acaccgccgg tccgacgcgg cccgacgggt
      901 ccgaggcatc gataaaataa aagattttat ttagtctcca gaaaaagggg ggaatgaaag
      961 accccacctg taggtttggc aagctagctt aagtaacgcc attttgcaag gcatggaaaa
     1021 atacataact gagaatagag aagttcagat caaggtcagg aacagatgga acagctgaat
     1081 atgggccaaa caggatatct gtggtaagca gttcctgccc cggctcaggg ccaagaacag
     1141 atggaacagc tgaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct
     1201 cagggccaag aacagatggt ccccagatgc ggtccagccc tcagcagttt ctagagaacc
     1261 atcagatgtt tccagggtgc cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa
     1321 ccaatcagtt cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct caataaaaga
     1381 gcccacaacc cctcactcgg ggcgccagtc ctccgattga ctgagtcgcc cgggtacccg
     1441 tgtatccaat aaaccctctt gcagttgcat ccgacttgtg gtctcgctgt tccttgggag
     1501 ggtctcctct gagtgattga ctacccgtca gcgggggtct ttcacatgca gcatgtatca
     1561 aaattaattt ggtttttttt cttaagtatt tacattaaat ggccatagtt gcattaatga
     1621 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     1681 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     1741 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     1801 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     1861 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     1921 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     1981 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2041 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2101 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2161 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     2221 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2281 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2341 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2401 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2461 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     2521 aggatcttca cctagatcct tttgcggccg caaatcaatc taaagtatat atgagtaaac
     2581 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     2641 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     2701 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     2761 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     2821 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     2881 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     2941 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     3001 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     3061 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     3121 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     3181 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     3241 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     3301 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     3361 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     3421 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     3481 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     3541 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgcagcct gaatatgggc
     3601 caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga acagatggaa
     3661 cagctgaata tgggccaaac aggatatctg tggtaagcag ttcctgcccc ggctcagggc
     3721 caagaacaga tggtccccag atgcggtcca gccctcagca gtttctagag aaccatcaga
     3781 tgtttccagg gtgccccaag gacctgaaat gaccctgtgc cttatttgaa ctaaccaatc
     3841 agttcgcttc tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa aagagcccac
     3901 aacccctcac tcggggcgcc agtcctccga ttgactgagt cgcccgggta cccgtgtatc
     3961 caataaaccc tcttgcagtt gcatccgact tgtggtctcg ctgttccttg ggagggtctc
     4021 ctctgagtga ttgactaccc gtcagcgggg gtctttcatt tgggggctcg tccgggatcg
     4081 ggagacccct gcccagggac caccgaccca ccaccgggag gcaagctggc cagcaactta
     4141 tctgtgtctg tccgattgtc tagtgtctat gactgatttt atgcgcctgc gtcggtacta
     4201 gttagctaac tagctctgta tctggcggac ccgtggtgga actgacgagt tctgaacacc
     4261 cggccgcaac cctgggagac gtcccaggga ctttgggggc cgtttttgtg gcccgacctg
     4321 aggaagggag tcgatgtgga atccgacccc gtcaggatat gtggttctgg taggagacga
     4381 gaacctaaaa cagttcccgc ctccgtctga atttttgctt tcggtttgga accgaagccg
     4441 cgcgtcttgt ctgctgcagc gctgcagcat cgttctgtgt tgtctctgtc tgactgtgtt
     4501 tctgtatttg tctgaaaatt agggccagac tgttaccact cccttaagtt tgaccttagg
     4561 tcactggaaa gatgtcgagc ggatcgctca caaccagtcg gtagatgtca agaagagacg
     4621 ttgggttacc ttctgctctg cagaatggcc aacctttaac gtcggatggc cgcgagacgg
     4681 cacctttaac cgagacctca tcacccaggt taagatcaag gtcttttcac ctggcccgca
     4741 tggacaccca gaccaggtcc cctacatcgt gacctgggaa gccttggctt ttgacccccc
     4801 tccctgggtc aagccctttg tacaccctaa gcctccgcct cctcttcctc catccgcccc
     4861 gtctctcccc cttgaacctc ctcgttcgac cccgcctcga tcctcccttt atccagccct
     4921 cactccttct ctaggcgccg gccggatccc agtgtggtgg tacgtaggaa ttcgccagca
     4981 cagtggtcga cgccgccacc atggattata aagatgacga cgataaggcg gcgctgagcg
     5041 gtggcggtgg tggcggcgcg gagccgggcc aggctctgtt caacggggac atggagcccg
     5101 aggccggcgc cggcgccggc gccgcggcct cttcggctgc ggaccctgcc attccggagg
     5161 aggtgtggaa tatcaaacaa atgattaagt tgacacagga acatatagag gccctattgg
     5221 acaaatttgg tggggagcat aatccaccat caatatatct ggaggcctat gaagaataca
     5281 ccagcaagct agatgcactc caacaaagag aacaacagtt attggaatct ctggggaacg
     5341 gaactgattt ttctgtttct agctctgcat caatggatac cgttacatct tcttcctctt
     5401 ctagcctttc agtgctacct tcatctcttt cagtttttca aaatcccaca gatgtggcac
     5461 ggagcaaccc caagtcacca caaaaaccta tcgttagagt cttcctgccc aacaaacaga
     5521 ggacagtggt acctgcaagg tgtggagtta cagtccgaga cagtctaaag aaagcactga
     5581 tgatgagagg tctaatccca gagtgctgtg ctgtttacag aattcaggat ggagagaaga
     5641 aaccaattgg ttgggacact gatatttcct ggcttactgg agaagaattg catgtggaag
     5701 tgttggagaa tgttccactt acaacacaca actttgtacg aaaaacgttt ttcaccttag
     5761 cattttgtga cttttgtcga aagctgcttt tccagggttt ccgctgtcaa acatgtggtt
     5821 ataaatttca ccagcgttgt agtacagaag ttccactgat gtgtgttaat tatgaccaac
     5881 ttgatttgct gtttgtctcc aagttctttg aacaccaccc aataccacag gaagaggcgt
     5941 ccttagcaga gactgcccta acatctggat catccccttc cgcacccgcc tcggactcta
     6001 ttgggcccca aattctcacc agtccgtctc cttcaaaatc cattccaatt ccacagccct
     6061 tccgaccagc agatgaagat catcgaaatc aatttgggca acgagaccga tcctcatcag
     6121 ctcccaatgt gcatataaac acaatagaac ctgtcaatat tgatgacttg attagagacc
     6181 aaggatttcg tggtgatgga ggatcaacca caggtttgtc tgctaccccc cctgcctcat
     6241 tacctggctc actaactaac gtgaaagcct tacagaaatc tccaggacct cagcgagaaa
     6301 ggaagtcatc ttcatcctca gaagacagga atcgaatgaa aacacttggt agacgggact
     6361 cgagtgatga ttgggagatt cctgatgggc agattacagt gggacaaaga attggatctg
     6421 gatcatttgg aacagtctac aagggaaagt ggcatggtga tgtggcagtg aaaatgttga
     6481 atgtgacagc acctacacct cagcagttac aagccttcaa aaatgaagta ggagtactca
     6541 ggaaaacacg acatgtgaat atcctactct tcatgggcta ttccacaaag ccacaactgg
     6601 ctattgttac ccagtggtgt gagggctcca gcttgtatca ccatctccat atcattgaga
     6661 ccaaatttga gatgatcaaa cttatagata ttgcacgaca gactgcacag ggcatggatt
     6721 acttacacgc caagtcaatc atccacagag acctcaagag taataatata tttcttcatg
     6781 aagacctcac agtaaaaata ggtgattttg gtctagctac agagaaatct cgatggagtg
     6841 ggtcccatca gtttgaacag ttgtctggat ccattttgtg gatggcacca gaagtcatca
     6901 gaatgcaaga taaaaatcca tacagctttc agtcagatgt atatgcattt ggaattgttc
     6961 tgtatgaatt gatgactgga cagttacctt attcaaacat caacaacagg gaccagataa
     7021 tttttatggt gggacgagga tacctgtctc cagatctcag taaggtacgg agtaactgtc
     7081 caaaagccat gaagagatta atggcagagt gcctcaaaaa gaaaagagat gagagaccac
     7141 tctttcccca aattctcgcc tctattgagc tgctggcccg ctcattgcca aaaattcacc
     7201 gcagtgcatc agaaccctcc ttgaatcggg ctggtttcca aacagagg
//
