Mapping 250K/500K SNP assay

36 GeneChip ® Mapping 500K Assay Manual
• Use only fresh reagents from the recommended vendors to help
eliminate changes in pH or the salt concentration of buffers.
• Properly store all enzyme reagents. Storage methods can profoundly
impact activity.
• When using reagents at the lab bench:
- Closely follow the protocol and ensure that enzymes are kept at
−20 o C until needed.
- Keep all master mixes and working solutions in chilled cooling
chambers as indicated.
- Properly chill essential equipment such as centrifuges, cooling
chambers and reagent coolers before use.
- Since enzyme activity is a function of temperature, ensure that all
temperature transitions are rapid and/or well-controlled to help
maintain consistency across samples.
Along with the enzymatic stages, lab instrumentation plays an
important role in WGSA. To aid in maintaining consistency across
samples and operators, all equipment should be well maintained and
calibrated including:
• The thermal cyclers used (pre-PCR, PCR, and post-PCR)
• GeneChip® Hybridization Oven 640
• GeneChip® Fluidics Station 450
• GeneChip® Scanner 3000 7G
• The UV spectrophotometer plate reader
• All multi-channel pipettes
Since WGSA involves a series of ordered stages, it follows that the
output of one stage directly impacts the performance of the subsequent
stage. For example, the quantity and purity of the DNA after
purification and normalization can affect the kinetics of the
Fragmentation Reagent during the subsequent fragmentation stage.
Variation in either the quantity or the relative purity of the DNA can
result in over- or under-fragmentation, which can lead to variability in
genotype call rates.
To efficiently process samples in a 96-well plate, it is essential to be
proficient with the use of 12-channel pipettes. Attempting to use a