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This high validation rate was achieved with minimal read depth and allele frequency filtering, and no additional filtering using often-employed properties, such as bidirectionality of reads and variant quality scores.
Nevertheless, even the minimal read depth of 78,000 allows for exceedingly precise determination of methylation levels.
While the level of concordance for mutational analysis using this panel compared to classical Sanger sequencing is excellent, we could 'only' multiplex 5 samples on a 316 chip to reach a minimal read depth of 500× per amplicon.
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In order to remove the contigs with minimal read depths that possibly resulted from so-called 'leaky' transcription [ 31], all assemblies were subjected to further modification as follows.
A minimum read depth of 3 reads is used to minimize false alignments of reads on the reference sequence.
Consequently, the minimum read depth parameter was fixed to 60.
The "-m" in genotypes means minimum reads depth for genotype.
We prepared ten IMR90 datasets whose sizes were 10 100%% of that of the full data, and used each of them to calculate domain boundaries under three different threshold values (i.e., minimal read depths) to calculate methylation levels.
The read depth minimum was set to 3 reads.
Additional file 6: Comparison of GeneRead V.1 and V.2 coverage at 100x minimum depth and mean read depth.
SNPs were called only for positions with a minimal mapping quality (-Q) of 20, and maximum read depth (-D) was set at 200.
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