Fig. 6

RALF1 regulates the flowering time. a qPCR analysis of the splicing efficiency of FLC introns 1 and 6 in the seedlings of wild-type and fer-4 mutants with or without RALF1 treatment. The samples were collected at ZT 4. The bar indicates the mean ± SD, and the experiments were independently repeated three times (ns. means no significant; *P < 0.05, **P < 0.01, Student’s t-test). b Different splicing variants of MAFs in Col-0 with or without RALF1 treatment as determined by semi-qPCR analysis. CS indicates constitutive splicing, and AS indicates alternative splicing. mRNA was isolated from 7-d-old seedlings and sample were collected at ZT 4. The results were repeated three times with similar result. c The flowering genotypes of wild-type and RALF1-OX plants under LD conditions. d Number of rosette leaves in WT (n = 15) and RALF1-OX (n = 15) under LD conditions. Data are presented as the mean ± SD, and the experiments were independently repeated three times with similar results (**P < 0.01; one-way ANOVA with Tukey’s test). e The flowering genotypes of wild-type, ralf1 and RALF1-RNAi grown under LD conditions. f Flowering time was measured by the number of rosette leaves at bolting. Error bars indicate the SD (n = 15). The experiments were independently repeated three times with similar results. Statistically significant differences are denoted with asterisks (*P < 0.05, **P < 0.01; one-way ANOVA with Tukey’s test). g Expression level of FLC detected by qPCR in 7-d-old Col-0, RALF1-OX and ralf1 mutant seedlings. Error bars indicate the SD of three technical replicates. Day and night are denoted by white and black bars, respectively. The experiments were performed at least four times. h Splicing efficiency of FLC intron 1 and intron 6 in the seedling of Col-0, RALF1-OX and ralf1 mutant. Seedlings were grown under LD conditions for 7 days and collect and ZT 12. The primer pairs F1/R1’ and F6’/R6 were used to detect the unspliced RNA for FLC introns 1 and 6, respectively. Primer pairs F1/R1 and F6/R6 were used to detect the spliced mRNA. The experiments were performed three times, the bar indicates the mean ± SD (*P < 0.05, **P < 0.01,one-way ANOVA with Tukey’s test)