TaqMan SNP Assay

Materials

• TaqMan® Genotyping Assay (supplied at 20X, 40X or 80X) (Applied Biosystems).
• TaqMan® Genotyping Master Mix (supplied at 2X)(Applied Biosystems).
• StepOnePlus Real-Time PCR System (Applied Biosystems).
• PC/Laptop with StepOnePlus Software installed.
• MicroAmp Fast 96-Well Plates.
• Optical Plate Covers.
• Nuclease-Free Water
• Sample Genomic DNA (extracted as described in Blood DNA Extraction)
• Postive control(s) (Minor allele) (optional, but recommended)

Protocol

1. Dilute gDNA samples to ng/ul.
2. Place gDNA, TaqMan® Genotyping Master Mix (40X), TaqMan® Genotyping Assay (2X) on ice.
3. Make up Reaction Master mix (see table below).

Table 1. Reaction Mix for full 96 well plate

Component	1X volume	96X Volume (+ 10% excess)
TaqMan® Genotyping Master Mix (2X)	5.00 ul	528.0 ul
TaqMan® Genotyping Assay (40X)	0.25 ul	26.4 ul
	5.25 ul	554.4 ul Reaction Mix (5.25 ul) added to each well (see Table 2.) Diluted gDNA samples each added to a single well (see Table 2.) Table 2. for full 96 well plate Component 1X volume 96X Volume (+ 10% excess) Reaction Master Mix 5.25 ul 554.4 ul gDNA samples 4.75 ul 501.6 ul 10 ul 1056 ul