Quantitative Measurement of Transposon Copy Number Using the Droplet Digital PCR

Abstract

Spontaneous proliferation of transposable elements contributes to genetic diversity at varying levels such as somatic mosaicism, genetic divergence in population, and genome evolution. Such genetic diversity is essential for plants’ adaptation to changing environment and serves as a valuable resource for crop improvement. Therefore, measuring the copy number variation of transposable elements with precision and efficiency is important to understand the extent of their proliferation. Droplet Digital PCR (ddPCR) is an accurate and sensitive technique that allows measurement of copy number variation of a transposon. Briefly, genomic DNA is extracted, digested, and partitioned into thousands of nanoliter-scale droplets. The TaqMan real-time PCR followed by the end-point fluorescence detection enables the quantitative measurement of copy number of template DNAs. Here in this chapter, we describe the step-by-step procedure of ddPCR using EVADE retrotransposon of Arabidopsis as an example.