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Extraction of Nucleic Acid Templates 27
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Extraction of Nucleic Acid Templates
John M. S. Bartlett
The following series of short technical descriptions covers the extraction of DNA
and RNA from various starting materials. We have gathered these together at the
beginning of this book to provide an easy reference. The polymerase chain reaction
(PCR) techniques described in the rest of this book are, in the main, worked laboratory
methods given detailed examples of the procedures used by the authors in their own
research. However, the aim is to provide the reader with a method that may be translated
into their own research; thus, although the description of ultrasensitive PCR focuses
on viral genomes and cancer, this method may of course be equally applied to DNA
from other sources. Rather than leaving the reader who is interested in applying this
technique to ancient DNA or DNA from bone, etc., to search through the various
chapters to find such a technique, we have collected these together for reference here.
PCR provides a simple method for the amplification and analysis of DNA; however,
for most applications involving PCR, the DNA (or cDNA for RNA methods) must be
in a reasonably pure state. Therefore, the first stage of any experimental procedure
involving PCR based technologies is the provision of a pure suspension of nucleic
acids, either RNA or DNA.
Extraction of nucleic acids is a fundamental precursor to almost all the techniques
described within this volume. Isolation of RNA and DNA from blood and
fresh tissues can be performed using a variety of techniques, which also form the
basis of methods of extraction of these substrates from other sources. The sensitivity
of PCR methods is now such that extraction of DNA and RNA from tissues fixed in
formaldehyde and buffered formalin is considered routine, and we are now able to
extract DNA from ancient tissues, feces, and many other sources. Indeed, in forensic
science, DNA fingerprinting from sources as diverse as residual saliva on food and
microscopic blood deposits is now possible! Indeed, the description of the extraction
of DNA/RNA alone could probably fill several major chapters. It has, however, not
proven desirable or feasible to be exhaustive in our approach to DNA/RNA extraction
protocols, and we have therefore restricted these to major methods in use in many
laboratories. Further references (1–7) that provide detailed reviews of methods for
nucleic acid extraction and some recommended web sites are listed in the reference
section.
From: Methods in Molecular Biology, Vol. 226: PCR Protocols, Second Edition
Edited by: J. M. S. Bartlett and D. Stirling © Humana Press Inc., Totowa, NJ
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