Abstract
We developed and modified a precise, rapid, and reproducible protocol isolating high‐quality RNA from tissues of multiple varieties of cassava plants (Manihot esculenta Crantz). The resulting method is suitable for use in mini, midi, and maxi preparations and rapidly achieves high total RNA yields (170–600 μg·g−1) using low‐cost chemicals and consumables and with minimal contamination from polysaccharides, polyphenols, proteins, and other secondary metabolites. In particular, A 260 : A 280 ratios were > 2.0 for RNA from various tissues, and all of the present RNA samples yielded ribosomal integrity number values of greater than six. The resulting high purity and quality of isolated RNA will facilitate downstream applications (quantitative reverse transcriptase‐polymerase chain reaction or RNA sequencing) in cassava molecular breeding.
Highlights
Babak Behnam1, Adriana Bohorquez-Chaux2, Oscar Fernando Castaneda-Mendez2, Hiroyuki Tsuji1, Manabu Ishitani2 and Luis Augusto Becerra Lopez-Lavalle2
Samples were collected from storage roots at times that corresponded with differential expression studies of beta-carotene contents in roots of cassava strains at 12 months of age
We extracted RNA from cassava leaves using TRIZOL reagent according to previously described methods [20]
Summary
An optimized isolation protocol yields high-quality RNA from cassava tissues (Manihot esculenta Crantz). We developed and modified a precise, rapid, and reproducible protocol isolating high-quality RNA from tissues of multiple varieties of cassava plants (Manihot esculenta Crantz). The resulting method is suitable for use in mini, midi, and maxi preparations and rapidly achieves high total RNA yields (170–600 lgÁgÀ1) using low-cost chemicals and consumables and with minimal contamination from polysaccharides, polyphenols, proteins, and other secondary metabolites. Several reagents and kits are commercially available for isolating RNA from plants, including TRIZOL from Sigma-Aldrich; RNeasy Plant Mini Kits from Qiagen; and RNAiso Plus from Takara These kits fail in some plant tissues and in species with high polysaccharide, polyphenol, protein, and other secondary metabolite contents [12]. Our high-quality RNA isolation protocol is simple, nontoxic, and produces sufficient quantities of high-quality RNA
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
