Since the next-generation sequencing (NGS) systems were invented and introduced to life science research about a decade ago, the NGS technology has extensively utilized in wide range of genomic, transcriptomic, and evolutionary studies. Compared with other eukaryotic species, the application of NGS technology in plant research reveals some challenges in sample preparation and data analysis due to some structural and physiological characteristics and genome complexity nature in plants. Hence, despite of the standard sample preparation and data process protocols widely used in high throughput transcriptomic analysis, we also describe the modified hot borate RNA extraction protocol specific for high quality and quantity plant total RNA isolation, and some comments and suggestions to achieve better assessments in the validation of RNA and library quality and data analysis.
Keywords: Differential gene expression analysis; Hot borate; Illumina; RNA-Seq; Transcriptome.