Abstract
Obtaining a sufficient quantity of high-quality, intact RNA is the first crucial step in its study by RNA sequencing on next-generation sequencing platforms or quantitative PCR. Different RNA extraction methods or commercial kits vary in yield and in the quality and integrity of the RNA obtained, which may affect the results of downstream applications. Often, these factors depend on the organism under study and nature of the sample. Therefore, the selection of an appropriate RNA isolation method is critical. In this study, we present the results of an evaluation of three different commercial kits for the isolation of total RNA from Erwinia amylovora in apple tissue as well as the usefulness of different kinds of Deoxyribonuclease I for DNA removal and kits for rRNA depletion. To our knowledge, this is the first report on the method of isolation of high-quality E.amylovora mRNA for RNA-seq.
Highlights
Erwinia amylovora is a polyphagous bacterium causing fire blight on apple, pear and over 130 other plant species belonging mainly to the Rosaceae family (Van der Zwet and Keil 1979)
E. amylovora strains are very homogeneous in terms of phenotypic and genetic features, they show variations in pathogenic ability (Cabrefiga and Montesinos 2005; Hevesi et al 2000; Puławska et al 2006; Sholberg et al 2001; Norelli et al 1986)
Idared/M.26 were inoculated with E. amylovora strain 650 in greenhouse conditions in the spring
Summary
Erwinia amylovora is a polyphagous bacterium causing fire blight on apple, pear and over 130 other plant species belonging mainly to the Rosaceae family (Van der Zwet and Keil 1979). This pathogen, which causes great economic losses in the areas of apple and pear tree cultivation worldwide, has been the subject of several phenotypic and genetic diversity studies. E. amylovora strains are very homogeneous in terms of phenotypic and genetic features, they show variations in pathogenic ability (Cabrefiga and Montesinos 2005; Hevesi et al 2000; Puławska et al 2006; Sholberg et al 2001; Norelli et al 1986)
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