These data indicate that the proportion of templates amplified varied between 26 61% of the total templates available.
In general, non-quantitative methods of methylation analysis should be avoided as they detect only the presence or absence of methylation regardless of the extent of methylation and will score a sample as methylated even if only a small proportion of templates are methylated.
The proportion of methylated templates was then quantified according to methodology previously described [ 111].
BPCR ought to be utilizable to assay the proportion of input templates with 3' ends in the region of sequence similarity.
Remarkably, a significant proportion of detected template-directed adenine and uracil additions in sequence reads derived from the 5'-arm is associated with miR-281.
The relative proportions of each template amplified were determined by using the IQ5 software (Bio-Rad), employing the ΔΔCt method [ 15, 33, 34] to compare gene expression data.
In order to find the best proportion of these three templates for obtaining the highest FoF1 activity, we tested various proportions of the template DNA mix, at first.
Technically, RNA analysis is more amenable than DNA analysis to broader repertoire coverage from a given amount of starting template, given the larger proportion of actual input TCR template molecules.
At CpG site U11, the proportion of densely methylated templates in FAST-1 overexpressing fibroblasts significantly increased from 8% in unaffected controls to 79% (P < 0.01) and 76% (P < 0.001) in FAST-1 B and FAST-1 C, respectively, similar to the levels found in FRDA fibroblasts (Fig. 6b).
A possible explanation of the lower amplification efficiency with LNA-spiked primers is that for short targets such as miRNAs the primers that are incorporated into the template during amplification will lead to a high proportion of LNA in the template that will decrease the efficiency of subsequent PCR cycles.
