Variation in the RAD51 gene and familial breast cancer

Breast Cancer Research

Table 1 RAD51 PCR conditions

Exon	PCR fragment	Forward primer	Reverse primer	Annealing temperature (°C)^a	Size (bp)	DHPLC temperature (°C)
1	1	GCAAGCGAGTAGAGAAGTGGA	AACTGCCGCTGAGCACTG	54	218	65
2	2	ATGGCCTTGGCTTTTCCTAA	GGCCCTGCCAGACATATTTA	54	391	57
3	3	TGGAACCAACTTCCCATCTC	TTCCCACTAATGCCTCCCTA	54	341	58
4	4	CTCTTCCCATTGCACACCTT	CACCTGGCCTTCCTCTATCTC	54	371	52, 57
5	5	TCTGATGAGCTCCAAGAACA	TGACATGGAAGGATTTTGAAG	51	344	58
6	6	AAGGGAATGCCTCCTTCCTA	CCAAACTAACCCTGGCAATC	54	314	59
7	7	CAAATTGCTCATCTGCCTG	TGAGGCACCGTTTAACAAGA	54	397	59
8	8	TGGTAAGGAAGGGACCAGAA	TGTGGCCATAGACACTCCAA	57	390	60
9	9	TCGTTATTTTGTGGGGGAAA	ACAGGGGAGAGGCATATCAA	54	447	58
10	10a	TTGGTGCTTTGGTCTGTGTC	ATACCCCTCCTCCAAAACCA	54	409	59
10	10b	CAGGAGACAGGTCAGTAGTCACA	AGGTTTGGCACAAGACTCCA	51	374	57
10	10c	TGATCTTGTGTAAGGGTTTGGTT	GCAATCTCGACTCACTGCAA	51	375	60
10	10d	GATAGCCTGAGGTGGGAGAA	TCTGCAAGTGGGACTTTCCT	54	319	60

^a'Touchdown' PCR amplification conditions were as follows: denaturation at 94°C for 10 minutes, followed by two cycles of 94°C for 30 seconds, 30 seconds at the fragment annealing temperature (T_A) + 6°, and 72°C for 30 seconds. The conditions remained the same for the rest of the PCR except for the annealing temperature, which consisted of two cycles at (T_A + 4°C), then two cycles at (T_A + 2°C) and, finally, 35 cycles at the T_A. A final extension step was conducted at 72°C for 7 minutes.

ISSN: 1465-542X