Extended Data Fig. 11. Functional dissection of the Srh1 locus.
(a) Light microscopy of short- and long-haired rachillae at Waddington developmental stage W8.5-9 using DAPI staining to visualize the nuclei. Size differences of nuclei in epidermal and trichome cells are very obvious. The shown micrographs are representative of a total of five individual spikes sampled on separate days. (b) Densitometric measurement of DNA content in epidermal and trichome cells of DAPI stained rachillae of genotypes Morex and Barke, respectively. While trichome cells in short-haired rachillae undergo only one cycle of endoreduplication, the cells in long haired trichomes show eight to sixteen-fold higher DNA contents than epidermal cells indicating three to four cycles of endoreduplication. (c) mRNA in situ hybridization of HvSRH1 in longitudinal spikelet sections of Bowman with anti-sense (left) and sense (right) probes. The blue arrow indicates the position of a rachilla hair. Representative micrographs of two independent experiments are shown. (d) Principal coordinate analysis of SNP array genotyping data of different barley genotypes. Etincel and its mutant srh1P63S cluster together, proving their isogenicity. (e) srh1 mutant discovery. FIND-IT screenings identified a mutant with short-fuzzy hairs (top) in the background of the long-haired cultivar Etincel (bottom). The mutants are a P63S non-synonymous sequence exchange. Scale bar - 1 mm. Wildtype and mutant spikes were inspected for the srh phenotype. Spikes showed either the short- or long-hair phenotype (#mutant seeds: 22, #wild type seeds: 21), respectively. Individual representative seeds wer chosen for micrographic documentation. (f) HvSRH1 transcript abundance in RNA sequencing data of rachilla tissue in Barke (BA, long-haired), Morex (MX, short-haired), Bowman (BW, long-haired) and a short-haired near-isogenic line of Bowman (BW-srh). Samples were taken at two developmental stages: rachilla hair initiation (RI) and elongation (RE). Abundance was measured as transcripts per million (TPM). Points stands for individual biological replicates (n = 3). Error bars show the mean and standard error.