Development of New Total RNA Isolation Method for Tissues With Rich Phenolic Compounds

Abstract

Background: RNAs to be used in transcriptome analysis must be of high quality and pure in order to ensure maximum representation of the expressed genes. RNA isolation is difficult in hazelnut tissues containing large amounts of secondary metabolite, phenolic compounds and the cell wall structure. Commonly used protocols for RNA isolation are those that require a lot of labor and time and also do not allow sufficient RNA isolation when applied to tissues rich in phenolic compounds. This study was aimed to develop an efficient method for isolation of total RNAs from bud of hazelnut to be used in RNA sequencing. Materials and methods: An optimized new method was successfully applied on three different hazelnuts genotypes (cakildak, Palaz, Tombul) and about 25 times higher amount of total RNAs per mg fresh tissues were obtained compared to classical CTAB method. Different methods have been tried for the isolation of RNA from hazelnut tissues and the determination of the quality of the obtained RNAs. Results: The quality and quantity of isolalated total RNAs were determined by spectrophotometer, electrophoresis and PCR. This success has been caught without any compromise of purity since A(260)/A(280) ratios ranged from 1.90 to 2.04 and A(260)/A(230) ratios were >2.0 in all purified RNAs. Conclusion: The total RNAs isolated with new protocol was found to be suitable for RNA sequencing and other molecular applications.