CRISPR/Cas9-based knockout reveals that the clock gene timeless is indispensable for regulating circadian behavioral rhythms in Bombyx mori  

作　　者：Moses Addo Nartey Xia Sun Sheng Qin Cheng-Xiang Hou Mu-Wang Li 

机构地区：[1]Jiangsu Key Laboratory of Sericultural Biology and Biotechnology,School of Biotechnology,Jiangsu University of Science and Technology,Zhenjiang,Jiangsu,China [2]The Key Laboratory of Silkworm and Mulberry Genetic Improvement,Ministry of Agriculture,Sericultural Research Institute,Chinese Academy of Agricultural Sciences,Zhenjiang,Jiangsu,China

出　　处：《Insect Science》2021年第5期1414-1425,共12页昆虫科学（英文版）

基　　金：the National Natural Sci-ence Foundation of China(grant No.31772523);the Earmarked fund for the China Agriculture Research Sys-tem(cars-18).

摘　　要：Circadian rhythms,which are ubiquitous and adaptive,occur across all species,from microbes to humans,in which they organize and modify behavior and physiology.timeless(im)is a canonical clock gene.The core composition of the Drosophila melanogaster endogenous circadian clock has been extensively investigated;however,in lepidopteran insects,including Bombyr mori,the mechanism is complicated and little is known regarding the participation of tim in the negative feedback loop responsible for behavioral activities.To arrive at a comprehensive understanding of the role of tim in the B.mori endogenous circadian clock,we exploited the clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9 gene editing system.We attermpted to elucidate the functions of tim in the circadian clock of B.mori using Bmtim mutants.The knockouts affected two circadian behavioral activities:adult emergence and embryo hatching rhythms.Quantitative real-time polymerase chain reaction results confirmed that tim-knockouts induced relative reductions in the expression levels,and thereby the oscillation amplitudes,of Bmper and Bmclk messenger RNAs during both the photophase and scotophase.Additionally,the daily rhythmic expression of Bmndbt was up-regulated in the photophase and downregulated in the scotophase in a tim-knockout.Our study reveals that tim is integral to the B.mori circadian clock and may be involved in regulating eclosion and hatching rhythms.

关 键 词：Bombyx mori circadian trhythm CRISPR/Cas9 ECLOSION HATCHING JKT_CYCLE TIMELESS 

分 类 号：Q965[生物学—昆虫学]