Fig. 3: Map-based cloning of BTS-R.

a Genetic map of the BTS-R locus region in Arabidopsis. Positions of the markers used for mapping are indicated. The number of recombinants is indicated in parentheses. Predicted genes are shown by arrows. b The distribution of ΔSNP-index on chromosome 2. The MutMap analysis shows a genomic region with the highest ΔSNP-index peak harboring the candidate mutation. ΔSNP-index is obtained by subtracting the SNP-index value of bts-2 bulk from that of bts-r bulk. Blue dots corresponding to the ΔSNP-index value of homozygous SNPs were identified between bts-r and bts-2 mutant genomes. The red line represents the sliding window average values of ΔSNP-index indices of 1 Mb interval with a 1 kb increment. The green line represents the screening threshold value (95% confidence interval). The confidence interval was calculated by the bootstrap method to determine whether ΔSNP-index was significant. The dashed line represents ΔSNP-index = 0.5. c The genomic structure of the BTS-R gene. White and gray boxes indicate untranslated regions (UTR) and exons, and lines between boxes indicate introns. The positions of two sgRNA:Cas9 targets of RH24, T-DNA insertions, and the nucleotide substitutions in bts-r are shown. d The protein structure of RH24. RH24 protein contains a DEAD-box and a helicase domain. The early stop codon generated by the C to T mutation in bts-r is shown by the arrow. e Genetic complementation of bts-r by RH24. Phenotypes of Col-0, bts-2, bts-r, pRH24:RH24 (bts-r), pRH24:RH24^DAAD (bts-r), rh24-1/bts-2, pRH24:RH24 (rh24-1/bts-2), and pRH24:RH24^DAAD (rh24-1/bts-2) are shown. The genes were driven by the native promoter of RH24. The plants were grown at 22 °C for 4 weeks. Bar = 1 cm.