Simple and efficient RNA extraction and gene analysis in vegetative organs of Japanese persimmon

Abstract

This examination for simple and efficient RNA extraction methods was conducted in vegetative organs of persimmon, including leaves, which contain inhibitory substances. We applied the result to physiological genetic analysis. Among the four extraction kits and a traditional SDS+PVP method, the combination of FastPure and Fruit-mate was the most suitable, resulting in maximum yield of 3.96 μg with high quality from 20 mg of persimmon mature leaves. Extraction was also possible from several organs other than leaves. The standard curve of quantitative-PCR (Q-PCR) using extracted RNA presented a high correlation coefficient (r²=0.99). Using this method, the activity of photosynthesis, which is an important role of leaves, was analyzed using DkrbcL as a target. The seasonal expression pattern of DkrbcL mRNA in leaves which was clearly demonstrated, indicates that this RNA extraction method is very important for the genetic analysis of persimmon vegetative organs.