Development and application of single pollen nucleus genotyping and virus tools for meiotic gene and recombination studies in barley (Hordeum vulgare L.)

Abstract

Meiotic recombination generates genetic diversity and is essential for plant breeding, but crossover (CO) formation is tightly regulated and unevenly distributed in crops such as barley. Whether mechanisms identified in Arabidopsis thaliana also apply to barley remains unclear. This work establishes two complementary tools to study and modify the CO landscape in barley (Hordeum vulgare L.): multiplex single nucleus pollen genotyping (SNPG) and Barley stripe mosaic virus-induced genome editing (BSMVIGE). SNPG enables rapid measurement of male CO rates and interference, detecting up to four-fold increases in HvRECQ4 mutants. BSMVIGE allows efficient targeted editing of meiosis-related genes, including HvASY1, HvZYP1, and HvSCEP3. Depletion of synaptonemal complex components reduced chiasmata and fertility, highlighting their essential role in barley CO formation.