Abstract: 【Objective】Oxya chinensis is an important agricultural pest. The objective of this study is to reveal the function of carboxylesterases (Ces) genes and select potential molecular target for pest management through conducting the bioinformatics and tissue-specific expression analysis of Ces genes from O. chinensis transcriptome. 【Method】 The cDNA sequences of Ces genes were searched using the keyword “carboxylesterase” against the O. chinensis transcriptome database. The Ces cDNA sequence assembling, the deduced amino acid sequence analysis and the open reading frame finding were performed by using bioinformatics methods. Phylogenetic tree of O. chinensis carboxylesterase with complete ORF was constructed using neighbor-joining method. The structure, calculated molecular mass, isoelectric points, signal peptide and potential N-glycosylation sites of O. chinensis Ces amino acid sequences were predicted using corresponding online softwares. Optimal reference genes in different tissues of 5th instar nymphs were screened and relative expressions in different tissues of 10 Ces genes were detected by reverse transcription- quantitative PCR (RT-qPCR).【Result】 A total of 180 putative Ces cDNA fragments were identified from O. chinensis transcriptome. Among them 28 full-length cDNAs containing complete ORFs were obtained. Phylogenetic tree of Ces protein sequences from O. chinensis and other insect species showed that 28 O. chinensis Ces protein sequences were distributed in four clades, including clade A with Orthopteran and part of Dipteran α-esterases (20 Ces protein sequences), clade D with integumental esterases (2), clade E with β-esterases (4), and clade F with nonlepidopteran JHEs (2). With the exception of three Ces proteins with alkaline isoelectric points (pI), all the remaining Ces proteins are acidic or slightly acidic with pI ranging from 4.38 to 6.84. All the Ces protein sequences have N-glycosylation site, N-terminal conserved cysteine residues, and oxyanion hole. Signal peptide prediction showed 19 of 28 have signal peptide. Twenty-one Ces protein sequences have conserved catalytic triad S-E-H, however, the remaining 7 Ces protein sequences showed different amino acid substitutions. GeNorm and Normfinder analysis showed that EF1α and RP49 were the optimal reference genes. RT-qPCR showed that 10 Ces genes were expressed in 7 different tissues, in which OcCesA1, OcCesA6, OcCesA12, OcCesA14, OcCesA18 and OcCesA19 mainly expressed in the midgut and gastric caecum; OcCesA6, OcCesA18, OcCesA19 still expressed in Malpighian tubules; OcCesA2 highly expressed in the hindgut. OcCesD1 had the highest expression in Malpighian tubules, followed by the fat body and integument. OcCesE1 had the highest expression in gastric caeca, OcCesF2 had the highest expression in midgut.【Conclusion】A total of 28 Ces cDNAs with complete ORFs were identified from the O. chinensis transcriptome database. These Ces cDNAs were clustered into four different clades in the phylogenetic analysis. Significant expansions of the Ces genes were found in clade A and highly expressed in detoxification, excretory organs, such as midgut, gastric caeca and Malpighian tubules. These results contributed to the further functional analysis of O. chinensis Ces and biological control of O. chinensis based on Ces gene molecular target.

Key words: Oxya chinensis, transcriptome, carboxylesterase gene, bioinformatics, tissue-specific expression