Nucleic Acid Extraction from Human Biological Samples
Nucleic acid isolation is often the starting point for all downstream experiments in biomedical research. It is therefore the most crucial step in any molecular technique. DNA and RNA extraction follow protocols with standardized reagents, many of which are available in quality-controlled commercial kits. Irrespective of the protocol, successful extraction of high-quality nucleic acid from biological tissues requires sufficient disruption of the tissue and cellular structures, denaturation of nucleoprotein complexes, inactivation of nucleases, and nucleic acid purification. These steps can be modified based on nucleic acid of interest and biological sample source. This chapter addresses DNA and RNA extraction from a variety of sample and tissue types, including saliva, and formalin-fixed, paraffin-embedded tissues, which are often archived in clinical pathology laboratories. Special considerations and common pitfalls of each protocol will also be discussed, as will nucleic acid quantitation techniques.
Key wordsAmniocyte Blood Bone marrow aspirate Buccal cells Cultured cells DNA extraction Formalin-fixed Paraffin-embedded (FFPE) tissue Fresh/frozen tissue Nucleic acids RNA Saliva
The authors wish to acknowledge all of the members of the UCLA Molecular Diagnostics Laboratory who contributed to the development and validation of these methods. We also thank the UCLA Department of Pathology and Laboratory Medicine for their generous support.
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