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. 2004 Jan;63(1):98–101. doi: 10.1136/ard.2003.006056

Figure 1 .

Figure 1

Fas promoter genotyping by haplotype specific polymerase chain reaction (PCR). Four haplotype specific PCR reactions (table 1) were used to amplify cis combinations of the Fas promoter -1377 and -670 polymorphisms. Genomic DNA was amplified in 13 µl reaction mixtures consisting of 67 mM Tris base pH 8.8; 16.6 mM ammonium sulphate; 2 mM magnesium chloride; 0.01% vol/vol Tween 20; 200 mM of each of dATP, dTTP, dGTP, and dCTP; 0.1–0.01 µg DNA; 3.4 mM haplotype-specific primers, 6.8 mM control primers; and 0.1875 units of Taq polymerase. Thermal cycling parameters were: 1 minute at 96°C; 5 cycles of 96°C for 25 seconds, 70°C for 45 seconds and 72°C for 45 seconds, followed by 21 cycles of 96°C for 25 seconds, 65°C for 50 seconds and 72°C for 45 seconds; followed by 4 cycles of 96°C for 25 seconds, 55°C for 60 seconds and 72°C for 120 seconds. PCR products were electrophoresed in 1.0% agarose gels containing 0.5 mg/ml ethidium bromide and visualised under ultraviolet illumination. Direction of electrophoresis is upward.