Large portion of essential genes is missed by screening either fly or beetle indicating unexpected diversity of insect gene function

Abstract

Most gene functions were detected by screens in very few model organisms but it has remained unclear how comprehensive these data are. Here, we expanded our RNAi screen in the red flour beetle Tribolium castaneum to cover more than half of the protein-coding genes and we compared the gene sets involved in several processes between beetle and fly. We find that around 50 % of the gene functions are detected in both species while the rest was found only in fly (~10% ) or beetle (~40%) reflecting both technical and biological differences. We conclude that work in complementary model systems is required to gain a comprehensive picture on gene functions documented by the annotation of novel GO terms for 96 genes studied here. The RNAi screening resources developed in this project, the expanding transgenic tool-kit and our large-scale functional data make T. castaneum an excellent model system in that endeavor.

Phenotypic Screen and Transcriptomics Approach Complement Each Other in Functional Genomics of Defensive Stink Gland Physiology

“…iBeetle: Large-scale genome-wide phenotypic screen iBeetle stands for a large-scale systemic RNAi-based screen using the red our beetle, T. castaneum, as a screening platform. Double-stranded RNA injections were performed in larval or pupal stages and the phenotypic effects scored at multiple levels such as cell biology, physiology, or embryonic and postembryonic development [36,37]. In the rst and second phase of iBeetle, together about 8500 gene models were screened for their function by systematic gene knock-down, which corresponds to slightly more than 50% of the currently annotated genes in this emerged model organism [33].…”

“…In the second phase of the iBeetle screen, which covered 3400 genes, double-stranded RNA injection of iBeetle fragments (https://ibeetle-base.uni-goettingen.de/) [38,39] was performed at a concentration of 1 µg/µl into pupae of the PIG-19 strain [41]. Stink gland analysis was carried out at the adult stage 21 days after injection [37]. For the second phase, an additional 73 genes were noted in iBeetle.Base [39] to potentially have a knock-down-mediated altered phenotype.…”

“…The functional genetic tools in T. castaneum include forward genetics based on insertional mutagenesis [28], transgene-based miss-expression systems [29,30], a fully annotated genome sequence [31][32][33], as well as systemic RNAi [34,35]. The e cient use of RNAi in this beetle has allowed the development of this reverse genetics approach into a forward genetics application to perform an unbiased genome-wide large-scale phenotypic screen (iBeetle screen) to identify gene functions in embryonic and postembryonic development as well as cell biology and physiology [36,37]. In this screen, the knock-down situations were besides other phenotypes systematically checked for phenotypes in the stink glands and methodically documented in the iBeete-Base [38,39].…”

Persistent parental RNAi in the beetleTribolium castaneuminvolves maternal transmission of long double-stranded RNA

“…As a well-established model system, the red flour beetle Tribolium castaneum has been at the forefront of research on the RNAi mechanism [1,10,12] and for diverse genetics studies [13]. It is an effective RNAi screening platform [14][15][16]. pRNAi in Tribolium is regularly used for phenotypic investigation of development and to test genetic interactions singly or globally, such as by RNA-seq after RNAi [17][18][19][20].…”