Programmed cell death assays for plants

Programmed cell death (PCD) is a critical part of normal development at all stages of the plant life cycle. This chapter discusses protocols for PCD assays. DNA degradation is an important diagnostic factor in programmed cell death, and cleavage of DNA in internucleosomal fragments of 180 bp is considered a hallmark of apoptosis. The ability to isolate undegraded high molecular weight DNA from viable tissue is a requirement for studying PCD-induced DNA degradation because it ensures that any DNA degradation observed is not an artifact of the isolation procedure. The barley aleurone layer is a model system to study hormonally induced PCD in plants. In this tissue, gibberellic acid (GA) induces PCD, whereas abscisic acid (ABA) prevents this program. The chapter describes a method for DNA isolation from barley aleurone cells that minimizes DNA degradation during the purification process. It has been shown to work well for high molecular weight DNA analysis in a variety of other tissue and cell preparations, for example, Arabidopsis thaliana leaves.