Single cell RNA-seq analysis of individual sperm cells to identify transcriptomic differences potentially involved in preferential fertilization.

ended

Human societies are sustained by flowering plants in different ways but especially as a key food supplier. However, it is known that increasing global temperatures strongly affect plant reproduction and the growing human population will require more food in short-middle term. Thus, to create new crop varieties more resistant to heat stress and with enhanced yield, a deeper understanding of the mechanism of plant sexual reproduction will be crucial for increasing the efficiency of plant breeding programs. Plant reproduction relies on a double fertilization process, in which two sperm cells from a pollen grain reach the ovule and one fertilize the egg cell while the other the central cell. In some species with dimorphic sperm cells a preferential fertilization of each sperm cell has been described, but for Arabidopsis thaliana as well as for many economically important crops with isomorphic sperm cells this question remains unclear. The main goal of this project is to identify by single cell RNA-seq analysis potential transcriptomic differences between the two Arabidopsis sperm cells. To do that, individual sperm cells from pollen tubes growing through female stylar transmitting tissue (semi in vivo) will be isolated by micromanipulation using a specially designed microfluidic chamber on an inverted microscope. The identification of differentially expressed genes would enable for the first time the creation of a sperm single-cell marker line to eventually explore a hypothetical preferential fertilization in Arabidopsis. These results can be adapted for plant breeding programs to develop impactful biotechnological tools, like haploid inducer lines with genetically modified sperm cells to specifically deliver components in the egg cell to induce in vivo gynogenesis. Additionally, the results will be a valuable scientific resource to address other topics beyond the preferential fertilization, like exploring hybridization barriers based on gamete recognition.