Protocol

Preparing DNA from Blood for Genotyping

¹Rutgers University Cell and DNA Repository, Department of Genetics, Piscataway, NJ 08854-8082, USA
²Bionomics Research and Technology Center, Environmental and Occupational Health Sciences Institute, University of Medicine and Dentistry of New Jersey--Robert Wood Johnson Medical School, Piscataway, NJ 08854-5635, USA
³Molecular Diagnostics Program, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA

⁴Corresponding author (sahota{at}biology.rutgers.edu)

INTRODUCTION

This protocol describes the extraction of genomic DNA from whole blood samples (fresh or frozen) and buffy coats. It provides methods for (1) large-scale extraction of DNA using either in-house or commercial (PUREGENE) reagents; (2) mid-scale extraction of DNA using the QIAamp DNA Blood Midi Kit (for 0.5-mL samples); and (3) small-scale extraction of DNA using the QIAamp DNA Blood Mini Kits (for 200-µL samples). Also included are methods for extracting DNA from blood samples that are compromised or clotted. Compromised blood samples include samples that spent more than 3 days in transit, samples that have been stored for more than 1 wk at 4°C, samples that have been stored at -20°C or -70°C, and samples that were collected incorrectly (e.g., not mixed properly). The procedures described here are not high throughput. Potential drawbacks of making them high-throughput are lower recoveries of DNA and possible variability in data quality, thus requiring additional quality assurance and control checks. DNA yields for these methods vary with the number of nucleated cells in the blood sample and the quality of the sample.