Screening and expression analysis of chitinase genes of Mythimna separata based on the transcriptome sequencing
Chitinase in insects is crucial for their growth and development and serves as an important target for biopesticides.This study aims to identify chitinase genes in armyworm Mythimna separata,a significant migratory pest.High-throughput sequencing technology was employed to perform transcriptome sequencing,sequence assembly,functional annotation,and differential gene expression 3 500 analysis on the midgut and epidermal tissues of armyworm.Chitinase gene sequences were identified from the transcriptome database,and the physicochemical properties such as cDNA length,protein molecular weight,amino acid sequence,isoelectric point,instability index,transmembrane structure,and protein domains of these genes were predicted.A phylogenetic tree of chitinases from armyworm and other insects was constructed using MEGA software,and the expression patterns of the genes at different tissues and developmental stages were verified by q-PCR.A total of 19.42 Gb of data were obtained from the transcriptomes sequence of the midgut and epidermis of armyworm,with 25 236 Unigenes annotated in COG,GO,KEGG,KOG,Pfam,Swissprot,eggNOG,and nr databases.Gene expression analysis revealed 3 137 differentially expressed genes between the midgut and epidermis,with 1 872 highly expressed genes in the midgut and 1 265 in the epidermis.9 chitinase genes were identified from the transcriptome data,of which 7 genes were reported at first time,with cDNA lengths ranging from 1 362 to 9 816 bp.SMART structure prediction indicated that chitinases contain one or more catalytic domains.The constructed phylogenetic tree divided insect chitinase genes into nine subfamilies.q-PCR results showed that MsCht2,MsCht5,MsCht6,MsCht7,and MsIDGF1 were highly expressed in the epidermis,while MsCht4,MsCht11,and MsChi-H were highly expressed in the midgut,consistent with the transcriptome data.Most chitinase genes had the highest expression levels during the pupal or prepupal stages,on the other hand,MsCht4 exhibited the highest expression level during the 5th instar stage,while its expression level was considerably low during the pupa stage.There were distinct differences in gene expression between the midgut and epidermis of armyworm,indicating potential functional differences among different chitinase genes.The study identified 7 novel chitinase genes,providing new targets for the biological control of armyworm.The findings lay a foundation for further investigation into the functions of chitinases in army worm.
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