Arbitrary single primer amplification of trace DNA substrates yields sequence content profiles that are discriminatory and reproducible

• Published in: Electrophoresis Vol. 33; no. 3; pp. 492 - 498
• Main Authors: Waters, James M., Eariss, Graham, Yeadon, P. Jane, Kirkbride, K. Paul, Burgoyne, Leigh A., Catcheside, David E. A.
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY-VCH Verlag 01.02.2012; WILEY‐VCH Verlag
• Subjects: Amplification; Arbitrary primer; DNA - analysis; DNA - chemistry; DNA - metabolism; DNA Primers - chemistry; DNA Primers - metabolism; Forensic; Forensic Sciences; Humans; Metagenome; Nucleic Acid Amplification Techniques - methods; Nucleic Acid Amplification Techniques - standards; Oligonucleotide Array Sequence Analysis - methods; Profiling; Reproducibility of Results; Sensitivity and Specificity; Sequence Analysis, DNA - methods; Sequence Analysis, DNA - standards; Sequencing; Soil
• ISSN: 0173-0835; 1522-2683
• DOI: 10.1002/elps.201100359

Single primer amplification is shown to yield a DNA profile that is reproducible when based on the sequence content of the amplicons rather than on the pattern of length polymorphism. The sequence‐based profile increases in reliability with increasing numbers of cycles of amplification. This process uses an arbitrarily chosen primer and a low initial annealing temperature in order to amplify sequences from the whole metagenome present in a sample that may contain only trace DNA, and a large number of cycles to select subsets of sequences based on variable amplification efficiency. Using arrays, we demonstrate the utility and limitations of this approach for profiling the large metagenomes typical of soils and the trace DNA present in drug seizures. We suggest that this type of profiling will be most effective once next‐generation sequencing and advanced sequence analysis becomes routine.