Sources of Human Genomic DNA tested at Affymetrix

• Blood
• Cell line
• Formalin-fixed paraffin-embedded tissue (success will depend on quality, purity and quantity)

Genomic DNA Extraction/Purification Methods tested at Affymetrix

1. SDS/ProK digestion, phenol-chloroform extraction, Microcon® or Centricon® (Millipore) ultrapurification and concentration.
2. QIAamp® DNA Blood Maxi Kit (QIAGEN).

Methods that include boiling or strong denaturants are not acceptable, because the DNA would be rendered single-stranded.

Dissolve/elute DNA in reduced EDTA TE buffer (10 mM Tris, pH 8.0, 0.1 mM EDTA, pH 8.0) (TEKnova T0223)

Whole Genome Amplified Genomic DNA protocol tested at Affymetrix

REPLI-g® Kit (QIAGEN)

• Quantify DNA by Picogreen assay before amplification
• Use 30ng of genomic DNA for amplification, no further purification is needed
• Other pre-amplification methods have not been tested by Affymetrix

Recommended DNA Cleanup Procedure

Use if the genomic DNA preparation is impure or suspected to contain inhibitors.

1. Add to 250 ng genomic DNA
   • 0.5 volumes of 7.5M NH₄OAc
   • 2.5 volumes of absolute ethanol (–20°C)
   • 0.5 μL of glycogen (5 mg/mL)
2. Vortex and incubate at –20°C for 1 hour
3. Centrifuge at 12,000 x g in a microcentrifuge at room temperature for 20 minutes
4. Remove supernatant and wash pellet with 0.5 mL of 80% ethanol
5. Centrifuge at 12,000 x g at room temperature for 5 minutes
6. Remove the 80% ethanol and repeat the 80% ethanol wash one more time
7. Re-suspend the pellet in reduced EDTA TE buffer (10 mM Tris, pH 8.0, 0.1 mM EDTA, pH 8.0) (TEKnova T0223)