|CAMPBELL, BENJAMIN - University Of Minnesota|
|HOFSTAD, ANNA - University Of Minnesota|
|SREEKANTA, SUMA - University Of Minnesota|
|FU, FENGLI - University Of Minnesota|
|KOONO, THOMAS - University Of Minnesota|
|VANCE, CARROLL - Retired ARS Employee|
|MUEHLBAUER, GARY - University Of Minnesota|
|STUPAR, ROBERT - University Of Minnesota|
Submitted to: Theoretical and Applied Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/29/2016
Publication Date: 6/9/2016
Citation: Campbell, B.W., Hofstad, A., Sreekanta, S., Fu, F., Koono, T., O'Rourke, J.A., Vance, C.P., Muehlbauer, G.J., Stupar, R. 2016. Fast neutron induced structural rearrangements at a soybean NAP1 locus result in gnarled trichomes. Theoretical and Applied Genetics. doi: 1007/S00122-016-2735-x.
Interpretive Summary: A soybean plant with mutant trichomes was identified from a mutant soybean population. Genetic analyses including genetic mapping, DNA array hybridizations, RNA-sequencing, and whole genome re-sequencing identified a region within a single gene (Glyma.20G019300) that contained two sequence deletions and multiple sequence re-arrangements. The closest Arabidopsis homolog to this gene is the Nck-associated protein 1 (NAP1) gene, which is required for proper trichome development. The function of this gene in soybean was confirmed by transforming known Arabidopsis mutant plants with the wild-type soybean gene, which restored normal trichome development. A second soybean mutant with the same phenotype was identified from a separate mutant population. Sequence analysis of Glyma.20G019300 in this plant revealed the protein coding sequence is shorter resulting in a mutated protein. The combination of experiments determined Glyma.20G019300 is required for proper trichome development in soybean.
Technical Abstract: A soybean (Glycine max (L.) Merr.) gnarled trichome mutant, exhibiting stunted trichomes compared to wild-type, was identified in a fast neutron mutant population. Genetic mapping using whole genome sequence-based bulked segregant analysis identified a 26.6 megabase interval on chromosome 20 that co-segregates with the phenotype. Comparative genomic hybridization analysis of the mutant indicated that the chromosome 20 interval included a small structural variant within the coding region of a soybean ortholog (Glyma.20G019300) of Arabidopsis Nck-Associated Protein 1 (NAP1), an important regulator of actin nucleation during trichome morphogenesis. Sequence analysis of the candidate allele revealed multiple rearrangements within the coding region, including two deletions (approximately 1-2 kb each), a translocation, and an inversion. Further analyses revealed that the mutated allele perfectly co-segregated with the phenotype, and a wild-type soybean NAP1 transgene functionally complemented the Arabidopsis nap1 mutant (grl-4). Additionally, mapping and exon sequencing of NAP1 in a spontaneous soybean gnarled trichome mutant (T31) identified a frame shift mutation resulting in a truncation of the coding region. These data indicate that the soybean NAP1 gene is essential for proper trichome development and that the soybean gnarled phenotype resulted from complex structural rearrangements induced by fast neutron mutagenesis at the NAP1 locus.