Yang et al (2015) describe their successful knockout of the orco coreceptor using TALENs in the Asian corn borer, Ostrinia furnacalis.
Moving genetic technologies into new insect systems requires the solution to a number of problems including the development of embryo collection and microinjection protocols as well as mating and screening strategies. These problems are always challenging and can often be substantially difficult. Yang et al were successful in their efforts to deal with all of these challenges.
Ostrinia furnacalis lay eggs in clusters, each a monolayer and this presents a challenge to any efforts to deliver genetic technologies to appropriately aged embryos. Yang et al’s solved this by not really worrying about it and simply left the clusters of eggs intact and only injected the eggs around the periphery and then just prior to hatching destroying the embryos that were not injected. Over 40% of the eggs injected hatched.
Yang et al used TALENs to knock out the orco odorant coreceptor. They delivered mRNA encoding a pair of TALENs to embryos no more than 6 hours old.
Females arising from these injected embryos were mated and after laying eggs were were genotyped for evidence of orco mutations. Embryos from these females were reared to adults, mated and their eggs collected. The females were genotyped to identify those that were mutants.
The authors report data on two pairs of TALENS and one resulted in some 70% mosaicism with over 60% of the families yielded mutants. See the paper for all of the details.
Of course, the real purpose of this paper was to assess the phenotype of orco knockouts and homozygotes were unresponsive to sex pheromone.
This paper is not only interesting because of its technical successes but also the interesting phenotypes associated with orco knockouts and is worth reading.
Yang, B., Fujii, T., Ishikawa, Y., Matsuo, T., Targeted mutagenesis of an odorant receptor co-receptor using TALEN in Ostrinia furnacalis, Insect Biochemistry and Molecular Biology (2016), doi:10.1016/j.ibmb.2015.12.003.