Abstract: 【Aim】 To screen out the most stably expressed reference genes in Lygus pratensis under specific conditions. 【Methods】 qRT-PCR was adopted to determine expression levels of 10 candidate reference genes including alpha-tubulin, beta-tubulin, actin, elongation factor, succinate dehydrogenase, ubiquitin, transcription initiation factor TFIID subunit, glutathione S-transferase, ribosomal protein L32, and TATA-box binding protein genes in six categories of samples of L. pratensis including different sex adults, different adult tissues, different developmental stages, adults with different lambdacyhalothrin resistance, and adults respectively exposed to different temperatures and different insecticides. BestKeeper, geNorm, NormFinder and RefFinder were adopted to analyze the mRNA expression stability of the candidate reference genes. 【Results】 According to the results of different analysis methods based on the effective qRT-PCR data, it was concluded that the most stably expressed reference gene was RPL32 for different adult tissues and adults of L. pratensis with different lambda-cyhalothrin resistance. The most stably expressed reference genes for the groups of adults exposed to different temperatures, different sex adults, adults exposed to different insecticides and different developmental stages were UBQ, SDHA, β-tubulin and TAF, respectively. Based on the comprehensive analysis from geNorm and RefFinder calculation, the best combinations of stably expressed reference genes for the groups of different adult tissues, different sex adults, different developmental stages, adults with different lambda-cyhalothrin resistance, and adults exposed to different temperatures and different insecticides were RPL32+TAF, SDHA+GST, TAF+GST+UBQ, RPL32+GST, UBQ+ACT+β-tubulin and β-tubulin+TAF+RPL32, respectively. 【Conclusions】 All the most stably expressed reference genes and their best combinations obtained from this study could be acceptable as reference genes for gene expression analysis in L. pratensis for future studies.

Key words:  Lygus pratensis, qRT-PCR, reference gene, gene expression analysis, expression stability