An efficient method for total RNA extraction from leaves of arboreal species from the Brazilian Cerrado

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.

摘要 鉴于目前针对木本植物（arboreal species），尤其是巴西塞拉多（Brazilian Cerrado）植被的RNA提取相关研究资料匮乏，本研究旨在针对该生物群区的多种本土植物物种，筛选适配的RNA提取方法。本次测试的方法包含以下四类：(i) TRIzol®试剂、(ii) 改良型TRIzol®试剂、(iii) CTAB缓冲液（CTAB buffer）、(iv) 改良型CTAB缓冲液，实验初始阶段以Xylopia aromatica与Piper arboreum的叶片样品为测试材料。后续选取实验效果最优的提取流程，从另外17种本土植物物种中成功提取得到纯化总RNA。基于A260/A280吸光度比值的检测结果，改良型CTAB法是针对上述木本物种进行总RNA提取的最优方案。在通过选定方法提取的总RNA中，11种受试物种有10种可经逆转录PCR（RT-PCR）扩增得到预期大小的片段，这证实该方法是获取高质量RNA以开展分子分析，以及检测侵染此类树木的病毒的理想选择。