Similar mapping percentages were achieved using another mapping tool, Bowtie2 [ 28] (data not shown).
The mapping percentages were lower where there was not clear orthology relationship, but in every case, mapping to pig was greater than to mouse, and the degree of homology was also greater.
Interestingly, when we used transcript instead of genome sequences as a reference for mapping the zebrafish data, mapping percentages were between 40%and49%9% vs. up to 80% for the genome (data not shown).
In contrast, Figure 1 shows that the read mapping percentage is also sample dependent, and this holds true for every gene model.
Similarly, mRNA-seq read-mapping percentages were significantly higher using the MacaM assembly than the rheMac2 assembly (mean 85.2% vs. 70.0%, p <0.0001. When we compared mRNA expression levels with one set of animals at two time points, we detected many more Differentially Expressed Genes (DEGs) with the MacM genome than with rheMac2.
Assessing imprinted regions for methylation status is an important check to ensure appropriate mapping (i.e., if percentages deviate significantly form an expected norm, data should be re-assessed), but a gold standard experiment to determine if methylation mapping and percentages are consistent with informatics analysis can also be done.
Other possible reasons for low mapping percentages might be reads that map either to invalid fragments, which have been removed from analysis, or to new fragments, created by mutations relative to the reference genome.
Percentages are calculated according to the total number of reliable reads (MAPQ > 1).
The percentages are based on occupancies of CO found from the Fo – Fc omit map.
2 The percentages are transmogrified.
