A molecular toolkit for superorganisms

Abstract

Social insects, such as ants, bees, wasps, and termites, draw biologists' attention due to their distinctive lifestyles. As experimental systems, they provide unique opportunities to study organismal differentiation, division of labor, longevity, and the evolution of development. Ants are particularly attractive because several ant species can be propagated in the laboratory. However, the same lifestyle that makes social insects interesting also hampers the use of molecular genetic techniques. Here, we summarize the efforts of the ant research community to surmount these hurdles and obtain novel mechanistic insight into the biology of social insects. We review current approaches and propose novel ones involving genomics, transcriptomics, chromatin and DNA methylation profiling, RNA interference (RNAi), and genome editing in ants and discuss future experimental strategies.

Keywords: CRISPR; RNAi; ants; epigenetics; single-cell sequencing; transgenesis.

Figure 1.. Experimental approaches in ant biology.

See table 1 for a summary of specific experimental techniques, their advantages and disadvantages.

Figure 2.. Comparison of experimental strategies to obtain stable and conditional mutants.

This figure uses H. saltator as an example but this is also applicable to other species. On the left, previously used strategy to obtain Orco mutants (see [80] for method details). Cas9 protein and gRNA are combined in vitro and then injected into embryos. The embryos are allowed to hatch on agar plates and the larvae are placed into colonies with nursing workers (not shown). All animals develop into workers. Isolation of adult workers induces transition into an egg laying queen substitute (“gamergate”). Eggs laid by unfertilized females develop into males. Genotyping is done by Sanger sequencing. In males, DNA for genotyping is extracted from a clipped wing. Females are sacrificed and genotyped after a cross. P0 and F1-F4 are parental and offspring generations, respectively. “X” stands for “crossed to”. In genotype notations, “+” is a wild-type allele, “−” is a mutant allele, and “0” underscores that males are haploid. On the right, proposed strategy to generate transgenic animals for conditional mutagenesis or other applications. Instead of Cas9-gRNA complexes, a mixture of mRNA that codes for a transposase (in this example, the piggyBac transposase) and a plasmid that contains the construct to be inserted are injected instead. Black arrowheads on the piggyBac plasmid in the upper right corner symbolize inverted terminal repeat sequences recognized by the transposase. “PBac” in the genotype notations is an integrant allele. Green eye color symbolizes the expression of Px3P::GFP as an example of a selection marker.