Abstract: Monochamus alternatus is the main vector of pine nematode disease. The establishment of an efficient RNAi system will be helpful for the study of the functional genome of M. alternatus. In this study, we designed the RNAi position of Laccase 2 gene of M. alternatus, prepared dsRNA and siRNA using in vitro transcription and chemical synthesis, respectively, and optimized their injection conditions and injection age, combined with phenotypic observation and RT-qPCR to verify the RNAi silencing effect. The results showed that pupae injected with 1 μg and 10 μg dsRNA showed deformities after eclosion, no pigmentation and were unable to perform normal activities, while those injected with 4 μg siRNA showed no significant difference from the control. RT-qPCR expression analysis revealed that the expression of Laccase 2 was higher in the pupal to adult stages and reached the highest in the old mature pupae. The expression of Laccase 2 was significantly suppressed by dsRNA injection on the first day of pupal stage, and the silencing efficiency could reach about 70%. The dsRNA of Laccase 2 was used to inhibit the expression of target genes in M. alternatus, and the ideal silencing effect could be achieved by injecting 1 μg at the pupal stage, while siRNA injection had no effect in M. alternatus. This study provide technical support for further research based on functional genomics and provide a new direction for the biological control of M. alternatus.