Comparison of genome copy number per plastid by DAPI-DNA staining and real-time quantitative PCR (qPCR) for different tissues. (A) Genome copy number per plastid from DAPI-DNA fluorescence and plastid size for individual plastids. Plastids from immature tissue (entire shoots from 13-day-old seedlings) and mature tissue (first and second leaves of 20-day-old plants) are compared. The sample sizes are 68 and 53, respectively. (B) DNA obtained from the same samples described in (A) was quantified by real-time qPCR, and the mean genome copy number per plastid is compared to that value determined by DAPI-DNA fluorescence. For both methods, the mean copy number per plastid for the immature tissue was significantly greater than the mature tissue (P < 0.0001). Chloroplasts from young first and second rosette leaves of 12-day-old plants also exhibit a large range in genomes per plastid (0.8 to 203) that becomes reduced in mature first and second rosette leaves of 26-day-old plants (0 to 82) as determined by DAPI-staining (not shown). The mean genome copy number is also significantly lower for these mature leaves (24 ± 3) than for these immature leaves (58 ± 7). The sample sizes are 46 and 62, respectively.