Genome-Wide Structural Profiling and Bulk Segregant Sequencing Resolve a NAP1 Mutation Underlying a Short Trichome Phenotype in Soybean

A soybean (Glycine max (L.) Merr.) short trichome mutant was identified from in a fast neutron population in the background ‘M92-220’. A population was developed to map the cause of this mutant using bulk segregant analysis with whole genome sequencing. A 3:1 wild-type:mutant segregation ratio in the F₂ population indicated that a single locus was responsible for the mutant phenotype. A single round of mapping using bulks of 50 F₂ individuals identified an interval on chromosome 20 that cosegregated with the phenotype. In this interval, array Comparative Genomic Hybridization (aCGH) indicated a 1.5kb deletion was present in Glyma.20G019300, the soybean homolog of the Arabidopsis AT2G35110, or NAP1, gene. The Arabidopsis gene was identified as a component of the WAVE protein complex that is involved in activating the ARP2/3 complex involved in actin nucleation. Further investigation of the fast neutron mutation revealed that not only was there one deletion in this interval, but also a translocation, two inversions, and a second deletion. PCR primers were designed to either amplify the wild-type sequence or span one of the mutant inversion junction points. These primers were found to perfectly cosegregated with the phenotypes of 121 F₃ individuals. Efforts are underway to compliment the candidate gene in a mutant background using genetic transformation. This study was conducted as a proof of concept for utilizing bulk segregant analysis by whole genome sequencing in combination with aCGH to map and clone fast neutron induced mutations in soybean.