CRISPR/Cas9 ‐mediated Tyrosine hydroxylase knockout resulting in larval lethality in Agrotis ipsilon

AbstractTyrosine hydroxylase (TH) is involved in insect melanin and the catecholamine biosynthesis pathway. TH as an enzyme catalyzing the conversion of tyrosine to 3,4 ‐dihydroxyphenylalanine is the first step reaction in the pathway. Although TH has been proven to affect the pigmentation of the epidermis and development in many insects, there is no report about physiological function of theTH gene inAgrotis ipsilon. Here we cloned theTH gene fromA. ipsilon. Semi ‐quantitative real‐time polymerase chain reaction (PCR) analysis showed thatAiTH was expressed at all development stages. Moreover, its high expression levels in the head and epidermis suggest that it is mainly related to pigment deposition and insect development. Then, we used the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR ‐associated protein 9 system to target theAiTH gene: deletion events were detected at the target sites. Compared with the control group, a few mutants with the phenomenon of narrowing in the egg shell and embryos can develop but cannot hatch; the other hatched embryos were seriously dehydrated after hatching and died within the first day. Quantitative real ‐time PCR analysis revealed thatTH was down ‐regulated inAiTH mutants. Here, our work demonstrated thatAiTH plays an important role in growth and development of newly hatched larvae; meanwhile, it would be a promising target to explore a control strategy forA. ipsilon.

Source: Insect Science - November 22, 2018 Category: Biology Authors: Yang Yang, Yao ‐Hui Wang, Xi‐En Chen, Di Tian, Xia Xu, Kai Li, Yong‐Ping Huang, Lin He Tags: ORIGINAL ARTICLE Source Type: research