High-quality RNA, cDNA, and derived EST libraries from grapevine (Vitis vinifera L.)

Abstract

The implementation and generation of unbiased results from RNA-based techniques in functional genomic studies requires the isolation of high-quality RNA. In grapevines, the yield and quality of RNA can be significantly reduced by contaminants such as polyphenols, polysaccharides, and proteins, which are abundant during different stages of berry and leaf development. We describe an optimized cetyltrimethylammonium bromide (CTAB)-based RNA extraction protocol that allows effective extraction of high-quality total RNA from a wide range of grapevine tissues, including difficult organs such as symptomatic, pathogen-infected leaves and mature berries. Total RNA extracted with this protocol was successfully used for cDNA library construction as well as reverse transcription-polymerase chain reaction (RT-PCR) amplification. This protocol can also be easily scaled to accommodate different amounts of tissue.