Figure 10. Analysis of the distribution of 27 nt RNAs on macronuclear nanochromosomes.

A. 27 nt RNA small RNAs are produced in equal numbers from both strands of the nanochromosome. For each full length nanochromosome with at least ten 26–28 nt small RNAs aligning to each strand, the total number of 26–28 nt small RNAs that map to each strand was plotted (left graph) or the number of distinct 26–28 nt small RNAs that map to each strand was plotted (right graph). Pearson R correlation values of 0.91 for all reads and 0.94 for distinct reads were obtained. This indicates a strong correlation of small RNA production from one strand of the nanochromosome with production of small RNAs from the other strand of the nanochromosome. Red curved lines represent 2 standard deviations from the mean; 95% of points would be expected to fall within these regions if there is a one-to-one correlation between the number of 26–28 nt small RNAs aligning to each strand of a nanochromosome (thin black line along the main diagonal). The blue dotted line indicates the actual lines within which 95% of the data points fall. B. There is a non-uniform distribution in the positioning of small RNAs on the nanochromosomes. For each position in each complete nanochromosome, the number of 26–28 nt small RNAs that start at that position in the mat24_06 sequencing library were determined. Then we determined the mean and standard deviation of coverage density on the nanochromosome. The coefficient of variation (standard deviation of coverage divided by the mean coverage) is plotted for each nanochromosome in the histogram. The peak of coefficient of variation at ∼6.0 indicates that the standard deviation is 6.0 times greater than the mean. This is highly indicative of a non-uniform distribution of small RNA coverage on the nanochromosomes.