title: zhuanjianlv-9
organism: Toona sinensis
description: Protocols: Young tender leaflets from apical tips and fully expanded leaflets from basal positions of the same compound leaf were collected from three Toona sinensis cultivars exhibiting distinct leaf colors: red, brown, and green. Tissues were immediately frozen in liquid nitrogen and stored at -80°C until RNA extraction. Toona sinensis plants were grown under natural field conditions in a cultivation site in Beijing, China, with standard agricultural practices. Samples were collected during the spring growing season when leaf flushing occurred. Total RNA was extracted using the TRIzol reagent (Invitrogen) according to the manufacturer's instructions. RNA concentration and purity were determined using a NanoDrop spectrophotometer (OD260/280 ratio), and integrity was assessed by agarose gel electrophoresis. Stranded RNA-seq libraries were constructed using the NEBNext Ultra II Directional RNA Library Prep Kit (New England Biolabs) following the standard protocol. Poly(A) selection was performed to enrich for mRNA. Libraries were amplified with 12 PCR cycles and size-selected for fragments around 300 bp. Library quality and concentration were validated using an Agilent Bioanalyzer.