A METHOD FOR EXTRACTING HIGH-QUALITY RNA FROM DIVERSE PLANTS FOR NEXT-GENERATION SEQUENCING AND GENE EXPRESSION ANALYSES

作者：Yockteng Roxana; Almeida Ana M R; Yee Stephen; Andre Thiago; Hill Colin; Specht Chelsea D^*

来源：Applications in Plant Sciences, 2013, 1(12): 1300070.

DOI：10.3732/apps.1300070

• cDNA library
• gene expression
• Illumina
• RNA-Seq
• RNA extraction
• transcriptome

Premise of the study: To study gene expression in plants, high-quality RNA must be extracted in quantities sufficient for subsequent cDNA library construction. Field-based collections are often limited in quantity and quality of tissue and are typically preserved in RNA later. Obtaining sufficient and high-quality yield from variously preserved samples is essential to studies of comparative biology. We present a protocol for the extraction of high-quality RNA from even the most recalcitrant plant tissues.
Methods and Results: Tissues from mosses, cycads, and angiosperm floral organs and leaves were preserved in RNA later or frozen fresh at -80 degrees C. Extractions were performed and quality was measured for yield and purity.
Conclusions: This protocol results in the extraction of high-quality RNA from a variety of plant tissues representing vascular and nonvascular plants. RNA was used for cDNA synthesis to generate libraries for next-generation sequencing and for expression studies using quantitative PCR (qPCR) and semiquantitative reverse transcription PCR (RT-PCR).

• 出版日期2013-12