Figure 5.

Reduction of CKB gene expression by amiR-CKBfam delays circadian period in transgenic plants. A, Period estimates of GI promoter-driven luciferase expression in transgenic lines harboring control amiRNA and amiR-CKBfam. Lines with the longest period among the amiRNA control transformants and the lines with the longest period among the amiR-CKBfam transformants are shown (n = 8 for control amiRNA and 12 for CKBfam amiRNA). Luciferase activity was determined in T3 homozygous seedlings. Transgenic lines were entrained under 12-h-light/12-h-dark cycles for 7 d, transferred to constant red light, and luminescence levels were measured every 2 h for 5 d. Period lengths were determined using 24 to 120 h of free running using the BRASS program. Each symbol represents one seedling, and the lines indicate the average period value (thick) ± 95% confidence interval (thin; n = 32). Similar results were obtained in two independent trials. B, Relative expression of CKB1 in transgenic lines harboring control amiRNA and amiR-CKBfam. Transgenic lines were entrained under 12-h-light/12-h-dark cycles for 10 d and harvested at ZT9. Gene expression was determined as described in Figure 3B. Data represent mean ± SEM (n = 2).