Protocols for extraction of total RNA from pecan nut (Carya illinoinensis [Wangenh.] K. Koch) embryo tissue

Abstract

Introduction: Gene expression studies require extraction protocols that allow obtaining high quality RNA, especially when working with tissues rich in polysaccharides, lipids and polyphenols such as pecan nut (Carya illinoinensis [Wangenh.] K. Koch) embryo tissue. Objective: To evaluate the efficiency of eight methods of total RNA extraction from pecan nut embryo tissue. Materials and methods: Eight total RNA extraction protocols based on TRI Reagent®, CTAB (hexadecyltrimethylammonium bromide) buffer and a commercial kit were evaluated. Total RNA yield and quality were determined by spectrophotometry (UV/visible). RNA viability and integrity were analyzed by RT-PCR using actin as a reference gene. Results and discussion: Extraction protocols based on TRI Reagent® provided high concentrations of total RNA, but with a high degree of contamination. The commercial kit was used to extract total RNA, but without the expected optimal purity. Finally, protocols based on CTAB buffer achieved total RNA yields of optimal quality. Conclusions: The quality of total RNA varies according to the efficiency of the method used. The CTAB 4 protocol represents an efficient alternative for the isolation of RNA from embryonic tissues of C. illinoinensis.