LOCUS       sequence-494400-        7365 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This plasmid expresses the N-terminally 3x-FLAG tagged BRCT domain 
            that contains the Y1845D mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7365)
  AUTHORS   Gracia B, Montes P, Gutierrez AM, Arun B, Karras GI
  TITLE     Protein-folding chaperones predict structure-function relationships 
            and cancer risk in BRCA1 mutation carriers.
  JOURNAL   Cell Rep. 2024 Feb 27;43(2):113803. doi: 
            10.1016/j.celrep.2024.113803. Epub 2024 Feb 17.
  PUBMED    38368609
REFERENCE   2  (bases 1 to 7365)
  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: "Cell Rep.";
            date: "2024-02-27"; volume: "43(2):113803. doi"; pages: " 
            10.1016/j.celrep.2024.113803. Epub 2024 Feb 17"
COMMENT     Sequence Label: 
            pKlab522-pcDNA3-3xFLAG-V5-BRCA1(1314-end)-Y1845D-STOP
FEATURES             Location/Qualifiers
     source          1..7365
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     815..839
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             908..973
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             992..1033
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     protein_bind    1031..1055
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2712..2736)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2787..2804)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2793..3017
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3063..3491
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3150..3169)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3360..3381
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3500..3520)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3505..3834
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3685..3820
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3747..3766
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3883..4908
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3883..4908
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    5038..5159
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5075..5094)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5129..5148
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5208..5224)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5208..5224)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5221..5243)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5232..5248
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5256..5262,5263..5280,5281..5286))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5301..5322
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5439..5456)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5610..6198)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5690..5709)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6369..7160,7161..7229))
                     /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     6992..7011
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7230..7334)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagatcg
      841 cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgactc tagaggatcc
      901 agccatggac tacaaagacc atgacggtga ttataaagat catgacatcg actacaagga
      961 tgacgatgac aagggctcgg gctcgacctc gggtaagcct atccctaacc ctctcctcgg
     1021 tctcgattct acaagtttgt acaaaaaagt tggcgatcct ttcttgattg gttcttccaa
     1081 acaaatgagg catcagtctg aaagccaggg agttggtctg agtgacaagg aattggtttc
     1141 agatgatgaa gaaagaggaa cgggcttgga agaaaataat caagaagagc aaagcatgga
     1201 ttcaaactta ggtgaagcag catctgggtg tgagagtgaa acaagcgtct ctgaagactg
     1261 ctcagggcta tcctctcaga gtgacatttt aaccactcag cagagggata ccatgcaaca
     1321 taacctgata aagctccagc aggaaatggc tgaactagaa gctgtgttag aacagcatgg
     1381 gagccagcct tctaacagct acccttccat cataagtgac tcctctgccc ttgaggacct
     1441 gcgaaatcca gaacaaagca catcagaaaa agcagtatta acttcacaga aaagtagtga
     1501 ataccctata agccagaatc cagaaggcct ttctgctgac aagtttgagg tgtctgcaga
     1561 tagttctacc agtaaaaata aagaaccagg agtggaaagg tcatcccctt ctaaatgccc
     1621 atcattagat gataggtggt acatgcacag ttgctctggg agtcttcaga atagaaacta
     1681 cccatctcaa gaggagctca ttaaggttgt tgatgtggag gagcaacagc tggaagagtc
     1741 tgggccacac gatttgacgg aaacatctta cttgccaagg caagatctag agggaacccc
     1801 ttacctggaa tctggaatca gcctcttctc tgatgaccct gaatctgatc cttctgaaga
     1861 cagagcccca gagtcagctc gtgttggcaa cataccatct tcaacctctg cattgaaagt
     1921 tccccaattg aaagttgcag aatctgccca gggtccagct gctgctcata ctactgatac
     1981 tgctgggtat aatgcaatgg aagaaagtgt gagcagggag aagccagaat tgacagcttc
     2041 aacagaaagg gtcaacaaaa gaatgtccat ggtggtgtct ggcctgaccc cagaagaatt
     2101 tatgctcgtg tacaagtttg ccagaaaaca ccacatcact ttaactaatc taattactga
     2161 agagactact catgttgtta tgaaaacaga tgctgagttt gtgtgtgaac ggacactgaa
     2221 atattttcta ggaattgcgg gaggaaaatg ggtagttagc tatttctggg tgacccagtc
     2281 tattaaagaa agaaaaatgc tgaatgagca tgattttgaa gtcagaggag atgtggtcaa
     2341 tggaagaaac caccaaggtc caaagcgagc aagagaatcc caggacagaa agatcttcag
     2401 ggggctagaa atctgttgct atgggccctt caccaacatg cccacagatc aactggaatg
     2461 gatggtacag ctgtgtggtg cttctgtggt gaaggagctt tcatcattca cccttggcac
     2521 aggtgtccac ccaattgtgg ttgtgcagcc agatgcctgg acagaggaca atggcttcca
     2581 tgcaattggg cagatgtgtg aggcacctgt ggtgacccga gagtgggtgt tggacagtgt
     2641 agcactcgat cagtgccagg agctggacac ctacctgata ccccagatcc cccacagcca
     2701 ctactgataa tacccaactt tcttgtacaa agtggttgct ctatagtaac tcgagtctag
     2761 agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     2821 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     2881 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     2941 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     3001 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc
     3061 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     3121 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     3181 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     3241 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     3301 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     3361 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     3421 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     3481 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     3541 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     3601 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     3661 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     3721 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct
     3781 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     3841 ccgggagctt gtatatccat tttcggatct gatcagcacg tgatgaaaaa gcctgaactc
     3901 accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg
     3961 cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat
     4021 gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac
     4081 tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc
     4141 ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc
     4201 gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat
     4261 cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca
     4321 tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg
     4381 gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag
     4441 gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg
     4501 gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa
     4561 tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg
     4621 cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg
     4681 ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca
     4741 gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt
     4801 acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc
     4861 gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagca cgtgctacga
     4921 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     4981 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     5041 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     5101 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     5161 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     5221 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     5281 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     5341 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     5401 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     5461 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     5521 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5581 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5641 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5701 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5761 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5821 tatctcagtt cggtgtaggt cgttcactcc aagctgggct gtgtgcacga accccccgtt
     5881 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5941 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     6001 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     6061 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     6121 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     6181 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     6241 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     6301 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     6361 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     6421 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     6481 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     6541 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     6601 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     6661 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     6721 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     6781 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6841 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6901 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6961 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     7021 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     7081 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     7141 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     7201 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     7261 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     7321 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
