Functional Studies On Genes Related To Fatty Acid Biosynthesis During The Diapause Of The Ladybird,Coccinella Septempunctata

The widely distributed seven-spotted ladybird Coccinella septempunctata is a excellent natural predator that preys on various types of aphids,commonly found in agricultural and forestry plants.It plays an important role in biological control and is widely used as a special commodity.It overstays winter or summer by facultative diapause as adult.By regulating the diapause process and combining with the mass rearing technology,a realistic approach can be provided to prolong the shelf life of natural enemy products,improve the utilization effect of natural enemy products,and promote the progress of natural enemy insect industry.C.septempuncata can accumulate lipids significantly during diapause,which is the energy source for growth and development during diapause and post-diapause.Comprehensive analysis of this lipid accumulation can reveal the detailed molecular mechanisms underlying diapause regulation.As an important part of lipid accumulation,fatty acid biosynthesis pathway plays an important role in maintaining homeostasis in insects.Some key genes act together and interact with each other.Acetyl-Co A carboxylase(ACC)and long-chain fatty acid-Co A ligase(ACSL)are the key enzymes in fatty acid biosynthesis.elongase of very-long-chain fatty acids(ELO),and 3-ketoacyl Co A reductase(KAR)play crucial roles in the entire long chain fatty acid elongation pathway.According to previous transcriptomic data,the expression levels of these four enzymes were significantly up-regulated during diapause.It is speculated that the above four enzymes play important regulatory roles in the fatty acid metabolism pathway of C.septempunctata,but whether these four key enzymes in lipid metabolism are involved in the regulation of insect diapause and their specific roles in fatty acid metabolism remain unclear.In this study,the full-length sequences of CsACC,CsACSL,CsELO and CsKAR genes were obtained from C.septempunctata.The expression patterns of diapause induction and post-diapause were analyzed,and the functions of these four enzymes during diapause of C.septempunctata were investigated using RNAi technique.It is mainly divided into three parts:1.Based on the transcriptomic data of C.septempunctata,the full-length sequences of CsACC,CsACSL,CsELO and CsKAR genes were cloned by RT-PCR and RACE.The CsACC gene is 2971 bp long,with an open reading frame of 6792 bp,encoding 2263 amino acids;The CsACSL gene is 3890 bp long,with an open reading frame of 2046 bp,encoding 681 amino acids;The CsELO gene is 2773 bp long,with an open reading frame of 909 bp,encoding 302 amino acids;and the CsKAR gene is 1091 bp long,with an open reading frame of 945 bp,encoding 314 amino acids.The homology distribution of the four genes was basically consistent with the evolutionary relationships among species,and the four target genes were all most closely related to the homeotic gene from Tribolium castaneum.2.The results of q PCR showed that CsACC,CsACSL,CsELO and CsKAR genes were specifically expressed at the normal development stage,different diapause induction stages and post-diapause,and the expression level in early diapause was significantly higher than that in other stages.The expression pattern were consistent with the variation trend of the target genes in the transcriptomic data of C.septempunctata,and with the changes of lipid content and triglyceride contents in different stages in previous studies.3.Using RNAi technique to interfere with CsACC,CsACSL,CsELO and CsKAR genes,ds RNA treatment effectively reduced the transcriptional level of target genes,and the expression levels of each gene were significantly lower than that of the control group.After ds RNA injection,the total lipid content of each treatment group was significantly decreased,the water content increased extremely,the supercooling point increased distinctly,and the cold resistance was decreased.The abdominal lipid accumulation of ladybird was significantly less than that of the control group,and the development of digestive tract was more mature.This study confirmed that these four enzymes are involved in lipid accumulation and play important regulatory roles in the fatty acid metabolism pathway in C.septempunctata.