Figure 2.

Outline of the procedure for measuring Htr2c RNA editing. Total RNA was extracted from the homogenized amygdala of the mice brain using the TRIzol method. Both RNA extraction and cDNA synthesis were performed using PrimeScript Hi‐Fide RT‐RCR Kit. Then, the prepared cDNA samples (n = 5 cDNA samples/group) were methylated by bisulfite treatment using Bisul‐Methylation Universal Kit. Then, PCR amplification of Htr2c was performed, with the used primers designed by PyroMark Assay Design 2.0 and synthesized by Hua Da Gene Company, as shown in Table 1. Subsequently, serial pyrosequencing measures were performed with the substrate mixture, enzyme mixture, and four types of dNTP added in the reaction system. Finally, pyrosequencing detector and Pyro Q‐CpG software were used to measure the frequency for A, B, D, and C/E editing sites