*2.2. RNA Preparation and Sequencing*

Seeds from the 750-1 (low-N-tolerant) and ZCFB (low-N-sensitive) pepper cultivars were germinated, and the seedlings were cultured under two different N conditions (sufficient-N and low-N) as described above. Four weeks later, the youngest fully expanded leaves and roots of the seedlings grown in sufficient-N solution and low-N solution were collected separately at 10:00–11:00 A.M., flash frozen in liquid N, and then stored at −80 ◦C. Samples were collected from 45 independent plants from three replicates (15 plants/replicates). Total RNA was extracted using an RNA Plant Kit (Aidlab Biotech, Beijing, China). The quantity and quality of total RNA was examined on a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Madison, WI, USA) and an Agilent 2100 Bioanalyzer (Waldbronn, Germany), respectively. cDNA libraries were constructed and sequenced on a BGISEQ-500 platform at BGI (Shenzhen, China).
