the RNA-seq from total RNA extraction, library construction, and
data analysis.2 Materials
Prepare all solutions using ultrapure water and analytical grade
reagents. Prepare and store all reagents at room temperature
(unless indicated otherwise).2.1 Total RNA
Extraction
- Liquid nitrogen.
- 70% ethanol.
- Tissue: keep the tissue in the liquid nitrogen until the proce-
dure is completed. - TRIzol Reagent (Invitrogen).
- DEPC-treated water (Ambion).
- Chloroform: trichloromethane.
- Isopropanol.
- Thermo Scientific NanoDrop 2000 spectrophotometer: RNA
quantification - Agilent 2100 Bioanalyzer system: RNA quality control.
Fig. 1mRNA library construction workflow for Illumina (from David Corney 2013)
16 Hong Zhang et al.