CRISPR gene editing is now possible in cockroaches

CRISPR cartoon in cockroaches. Credit: Shirai et al./Cell Reports Methods

According to an article published in the journal Cell report methods by Cell Press on May 16^and, 2022, researchers developed a CRISPR-Cas9 technique to enable gene editing in cockroaches. The simple and effective “direct parental” (DIPA-CRISPR) CRISPR procedure involves injecting materials into adult females where the eggs are developing rather than into the embryos themselves.

“In a sense, insect researchers have been freed from the embarrassment of egg injections,” says the study’s lead author, Takaaki Daimon, of Kyoto University. “We can now edit insect genomes more freely and at will. In principle, this method should work for more than 90% of insect species.

Current insect gene-editing approaches typically require microinjection of materials into early embryos, severely limiting its application to many species. For example, previous studies have failed to genetically manipulate cockroaches due to their unique reproductive system. Moreover, insect gene editing often requires expensive equipment, species-specific experimental setup, and highly trained laboratory personnel. “These problems with conventional methods have plagued researchers who want to perform genome editing on a wide variety of insect species,” says Daimon.

To overcome these limitations, Daimon and collaborators injected Cas9 ribonucleoproteins (RNPs) into the main body cavity of adult female cockroaches to introduce inherited mutations into developing eggs. The results demonstrated that the efficiency of gene editing – the proportion of edited individuals out of the total number of hatched individuals – could be as high as 22%. In the red flour beetle, DIPA-CRISPR achieved an efficiency of more than 50%. Additionally, researchers have generated beetle knockin genes by co-injecting single-stranded oligonucleotides and Cas9 RNPs, but the efficiency is low and needs further improvement.

The successful application of DIPA-CRISPR in two evolutionarily distant species demonstrates its potential for large-scale use. But the approach is not directly applicable to all insect species, including fruit flies. Additionally, experiments have shown that the most critical parameter for success is the stage of adult females injected. Therefore, DIPA-CRISPR requires a good knowledge of ovarian development. This can be difficult in some species, given the various life cycles and reproductive strategies of insects.

Despite these limitations, DIPA-CRISPR is accessible, highly practical, and could be easily implemented in laboratories, extending the application of gene editing to a wide diversity of model and non-model insect species. The technique requires minimal equipment for injection in adults, and only two components – the Cas9 protein and the unique guide[{” attribute=””>RNA—greatly simplifying procedures for gene editing. Moreover, commercially available, standard Cas9 can be used for adult injection, eliminating the need for time-consuming custom engineering of the protein.

“By improving the DIPA-CRISPR method and making it even more efficient and versatile, we may be able to enable genome editing in almost all of the more than 1.5 million species of insects, opening up a future in which we can fully utilize the amazing biological functions of insects,” Daimon says. “In principle, it may be also possible that other arthropods could be genome edited using a similar approach. These include agricultural and medical pests such as mites and ticks, and important fishery resources such as shrimp and crabs.”

Reference: “DIPA-CRISPR is a simple and accessible method for insect gene editing” by Yu Shirai, Maria-Dolors Piulachs, Xavier Belles and Takaaki Daimon, 16 May 2022, Cell Reports Methods.
DOI: 10.1016/j.crmeth.2022.100215

This work was supported by funding from JSPS KAKENHI, JSPS Open Partnership Joint Research Projects, Spanish Ministry of Innovation and Competitiveness, and CSIC-Spain, and in part by Cabinet Office, Government of Japan, Cross-ministerial Moonshot Agriculture, Forestry and Fisheries Research and Development Program.