Analysis of Copy Number Variation by Pyrosequencing® Using Paralogous Sequences.

: The determination of gene copy numbers of highly similar genes is difficult with conventional PCR-based methods. However, by amplification of similar genes in the same PCR reaction followed by Pyrosequencing(®), one may distinguish the genes based on a single-nucleotide difference. The ratio between the peak heights of gene-specific nucleotides obtained in the Pyrosequencing reaction may thereby be used to calculate the relative copy numbers of target genes. This method is easy and cost effective compared to other methods, and allows for the determination of copy numbers of genes that were previously difficult to achieve.