Mapping 250K/500K SNP assay

114 GeneChip ® Mapping 500K Assay Manual
Stage 9: Target Hybridization
ABOUT THIS STAGE
During this stage, each sample is loaded onto either a GeneChip®
Human Mapping 250K Sty Array or a 250K Nsp Array. Three
methods for performing this stage are presented.
• Method 1 — Using a GeneAmp® PCR System 9700
Requires the use of a GeneAmp® PCR System 9700 located adjacent
to the hybridization ovens. Samples are on a 96-well reaction plate.
See Method 1 — Using a GeneAmp® PCR System 9700 on page 124.
• Method 2 — Using an Applied Biosystems 2720 Thermal Cycler or
an MJ Tetrad PTC-225 Thermal Cycler.
Requires the use of an Applied Biosystems 2720 Thermal Cycler or
an MJ Tetrad PTC-225 Thermal Cycler located adjacent to the
hybridization ovens. Samples are on a 96-well reaction plate. See
Method 2 — Using an Applied Biosystems 2720 Thermal Cycler or an MJ
Tetrad PTC-225 Thermal Cycler on page 127.
• Method 3 — Using Heat Blocks
Requires the use of two heat blocks and Eppendorf tubes, one per
sample. See Hybridizing Samples Using Heat Blocks on page 130.
First, you will prepare a Hybridization Master Mix and add the mix to
each sample. Then, based on the method you are using, you will
denature the samples on a thermal cycler (methods 1 and 2) or on a
heat block (method 3).
After denaturation, you will load each sample onto the appropriate
GeneChip Human Mapping 250K Array (Nsp or Sty) – one sample
per array. The arrays are then placed into a hybridization oven that has
been preheated to 49 °C. Samples are left to hybridize for 16 to
18 hours.
Two operators are required for all of the methods.