Patient clinical characteristics and family history.

<p>Patient clinical characteristics and family history.</p

Patient 3 pedigree.

<p>This pedigree represents the family history of cancer in patient number 3. The proband, or original patient, is denoted by the arrow. The type of cancer each relative has a history of is denoted below their pedigree symbol. If a relative had genetic testing and was found to carry the known familial <i>BRCA1</i> mutation, this is also denoted below their pedigree symbol. If a relative tested negative for the known familial <i>BRCA1</i>mutation, this is also denoted below their pedigree symbol. The proband had full gene sequencing and deletion/duplication analysis of the <i>BRCA1/2</i> genes; therefore, ‘<i>BRCA1/2-</i>‘ is listed below their pedigree symbol. The proband was also found to carry a variant of uncertain significance in <i>MSH2</i>.</p

<i>BRCA</i> phenocopy hypothesis.

<p>A. Maternal-fetal microchimerism. B. Tetragametic chimerism. C. A woman with <i>BRCA</i>-mutant chimeric cells.</p

Molecular testing in tumor tissue.

<p><b>A.</b> The <i>BRCA1</i> 187del AG deletion heterozygote (families 1 and 5) is detected as an n-2 product by capillary electrophoresis (left) and by an indicative peak pattern by pyrosequencing (top right). Neither the deletion product nor the mutant peak pattern was detected in the patient tumors (bottom panels). <b>B.</b> The heterozygote detected as a 163 bp product by gel electrophoresis. The synthetic oligomer carrying the mutation confirmed the detection of the mutation by mutation sequence specific primers. This band is not present in the negative control nor family 8 DNA (left four lanes). The 137 bp band specific to the normal A allele was detected by primers specific to that allele in the patient sample. Tumor DNA tested for the familial mutation 5215G>A gave similar results (not shown). <b>C.</b> The <i>BRCA2</i> 8107 A→T mutation was tested by sequence-specific PCR. The 92 bp product (T allele, positive) is not present in the patient’s tissue where only the A allele (80bp) is observed. Reagent blanks for the A and T allele primer sets (Bl A, Bl T) are shown. <b>D.</b> <i>BRCA1</i> 3109 insAA (left), and <i>BRCA2</i> 6794 insA (patients 10 and 11, respectively; right) mutation analysis by PCR-capillary electrophoresis. Amplified products from DNA without (negative control, top) and with (positive control, middle panels) demonstrate the expected right shift in migration for the <i>BRCA1</i> 3109 insAA n+2 product (76 bp) and the <i>BRCA2</i> 6794 insA n+1 product (82 bp)(bottom panels). Patient samples show an unexpected left shift (n-1) product.</p

Mutation primer sequences.

<p>Primers used to detect point mutations by sequence specific PCR, Inner primers end on the indicated variant bases. The size of the resulting product of the extended inner primer and its opposite outer primer will indicate the mutation status.</p