An evaluation of methods of blood preservation for RT-PCR from endangered species

Abstract

The study of conservation genetics traditionally uses an array of DNA based methods (Lambert and Millar 1995; Sunnucks, 2000). However, isolation of RNA may also be required, particularly in studies of genetic variation and disease where analysis of major histocompatibility complex (MHC) gene expression is important (Edwards and Potts 1996). Usually such studies use spleen or liver snap-frozen in liquid nitrogen for the isolation of high-quality RNA and the subsequent construction of cDNA libraries (e.g. Vincek et al. 1995). However this approach is not feasible for studies of endangered species, as often only small samples will be available, and a nonlethal method of sampling, such as from blood, is required. Several methods for the preservation of blood for DNA extraction exist (Carter 2000). In this study, we have tested these methods for their suitability for RNA extraction, then used an RTPCR based approach to isolate expressed class II B MHC sequences from the endangered Chatham Island black robin (Petroica traversi), from which only small amounts of blood (less than 200 匠per bird) can be sampled.