Transcriptome And Differential Expression Of Musca Domestica Larvae Under Temperature Stress

Housefly Musca domestica is a kind of temperature-changing animal.It has been proved that changes in environmental temperature will cause significant differences in morphology,in vivo protein and nucleic acid of housefly larvae,but the difference in transcriptome levels under temperature stress has not been reported.In this study,we used the Illumina HiSeq 4000 sequencing platform to perform transcriptome sequencing of 3rd-day housefly larvae cultured at 28°C at normal temperature,at temperature stress 18°C and 38°C,compared gene expression differences of housefly larvae under temperature stress.Expression of genes for functional annotation and metabolic pathway analysis.At the same time,8 of these genes were analyzed by real-time quantitative PCR to further verify the results of transcriptome sequencing.The main findings are as follows:45.03 Mb clean reads were obtained through transcriptome sequencing.43.570 unigenes were obtained after assembly and redundancy.Unigenes were compared to seven functional databases,and 33,789 unigenes had annotation information,which further enriched the transcriptome data of Musca domestica.This study provided basic data for studying the differences in transcriptional levels of housefly larvae under temperature stress.Using the transcriptome sequencing results as reference genes,the gene expression profiles of 3rd-day housefly larvae under temperature stress at 18°C(Md18)and 38°C(Md38)were compared with those at normal temperature 28°C(Md28).The expression of 10,863 and 11,606 genes in housefly larvae cultured at 18 °C and 38 °C changed significantly,among which 6,460 genes were significantly different in housefly larvae cultured at 18 °C and 38 °C.The differentially expressed genes were used to analyze the significant enrichment of GO functions.The results showed that in the classification of cell components,the three subtypes with prominently different proportions of Md18 and Md38 differentially expressed genes were cell,cell part and intracellular;in the molecular function classification,the most prominently expressed genes are binding and catalytic activities;in cell process classification,the proportion of differentially expressed genes in Md18 and Md38 were prominent.Both are cellular process,metabolic process,and single-organism process.The results show that temperature stress has a significant effect on the metabolic activity,apoptosis,proliferation and differentiation,and molecular binding of Musca domestica larvae.Eight differentially expressed genes(three heat shock protein genes(Unigene15829,Unigene31003,Unigene26272),three GST genes(CL1217.Contig3,Unigene25997,CL2294.Contig1)and two cytochrome P450 genes were selected from analysis of the expression profile results.CL914.Contig1,Unigene11717 validated the results of the expression profile.The results showed that 8 genes were up-regulated in Md18 and Md38 to varying degrees,which were consistent with the results of the expression profile,to indicate that the expression profile sequencing analysis results are more reliable,and this eight genes of the housefly larvae played a positive regulatory role in adapting to the ambient temperature.