Abstract
Abstract Considering the lack of information on RNA extraction from arboreal species, specially from the Brazilian Cerrado, the aim of this study was to test RNA extraction methods for a wide variety of native plant species from this biome. The methods tested consisted of: (i) TRIzol® reagent, (ii) TRIzol® reagent with modifications, (iii) CTAB buffer, and (iv) Modified CTAB buffer, initially for leaf samples of Xylopia aromatica and Piper arboreum. Later the procedure with the best results was used to obtain purified RNA from 17 other native species. Based on A260/A280 absorbance ratio the Modified CTAB method was the best for total RNA extraction for those woody species. Ten out of eleven species tested through RT-PCR generated fragments of the expected size from the total RNA extracted by the selected method, confirming it as the best option to obtain high-quality RNA for molecular analyses and for use in the detection of viruses infecting these tree species.
Highlights
Isolation of good quality RNA or DNA is the first step in studies related to plant molecular biology
Just a few total RNA extraction methods have proven to be efficient in tree species, such as Quercus robur L. (Kiefer et al 2000), Hevea brasiliensis (Willd. ex A.Juss.) Müll.Arg. (Deng et al 2012) and Neolamarckia cadamba (Roxb.) Bosser (Ouyang et al 2014)
Considering the A260/A280 absorbance values for P. arboreum, the values of Methods 1 and 2 were often below 1.5, which is below recommended for RNA quality (Glasel 1995)
Summary
Isolation of good quality RNA or DNA is the first step in studies related to plant molecular biology. Transcriptome studies of tree species help understand numerous plant processes under different conditions, such as gene expression associated with desirable agronomic characteristics (Deng et al 2016; Ouyang et al 2016; Shiraishi et al 2016) related or not to physiological processes such as flowering (Liu et al 2016) and wood generation (Wang et al 2016). These studies are dependent on obtaining relatively large amount of high-quality RNA. There is some difficulty in studying these plants at the molecular level, despite the fact that some species show desirable characteristics in different areas, including medical interest, production of antimicrobial compounds (Calixto-Júnior et al 2016; Correia et al 2016), and allelopathic potential in weed control (Candido et al 2016)
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