Fast neutron-induced structural rearrangements at a soybean NAP1 locus result in gnarled trichomes

Benjamin W. Campbell, Anna N. Hofstad, Suma Sreekanta, Fengli Fu, Thomas J. Y. Kono, Jamie A. O’Rourke, Carroll P. Vance, Gary J. Muehlbauer, Robert M. Stupar

Abstract

Key message

Three adjacent and distinct sequence rearrangements were identified at a NAP1 locus in a soybean mutant. Genetic dissection and validation revealed the function of this gene in soybean trichome development.

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 sequencing-based bulked segregant analysis identified a 26.6 megabase interval on chromosome 20 that co-segregated 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), a 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 mutant allele perfectly co-segregated with the phenotype, and a wild-type soybean NAP1 transgene functionally complemented an Arabidopsis nap1 mutant. In addition, 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 show the utility of the soybean fast neutron population for forward genetic approaches for identifying genes.

See: http://link.springer.com/article/10.1007/s00122-016-2735-x

Theoretical and Applied Genetics; September 2016; Volume 129, Issue 9, pp 1725–1738

Fig. 1

Phenotype and coarse genetic mapping of the gnarled trichome mutant. a Phenotypes of mutant (left) and wild-type (right) shoots. SEM leaf trichome images of the gnarled mutant R55C01 (b) and the wild-type line ‘M92-220’ (c). The mutant trichomes are wide, short, flaccid, and lay on the surface of the leaf or stem, as compared to the wild-type trichomes (narrow, long, straight, and project outward from the leaf or stem). Scale bars in b and c are 1 mm. d BSA-WGS allele frequencies were calculated for F₂ bulked samples that consisted of 50 mutant (red lines) and 50 wild-type (blue line) individuals. The allele frequencies were calculated as the proportion of reads containing the wild-type parental SNP (i.e. SNPs that match the wild-type parent ‘Noir 1’) at each position for over 16,000 polymorphic SoySNP50K positions. The allelic frequencies are shown as the average value across a 21 SNP sliding window. The obvious spread in allele frequencies indicates that the causative locus is located on chromosome 20 (color figure online)