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Currently, we analyze four samples together in a single Illumina HiSeq2500 lane in order to maintain a minimum read count of 20 million per sample.
A fold change ±1.5 with a minimum read count of ≥10 were used to filter the differentially expressed miRNA.
This result held when the minimum read count was increased to 200 or 500 reads per sex.
All unique sequence reads with a minimum read count of 10 were aligned with precursor chicken miRNA sequences from miRBase version 13.0 [ 19- 22].
Using the matched L2-poly(A) samples, a pseudogene was called expressed if it passed a minimum array intensity threshold of 100 or minimum read count of 1.
- Minimum read count for SNP call: Minimum number of non-reference allele counts required for a SNP to be considered detected.
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Therefore, we restricted our analysis to sequences with minimum read counts of 100.
Genotypes were scored only if a minimum total read count of 8 was met.
Using this "minimum 50" read count criterion, 729 IORs were identified as possibly containing novel sRNAs.
Novel predicted miRNA loci with an identified miRNA* sequence and a minimum miRNA read count of 5, as identified using the UEA Plant sRNA toolkit miRCat tool.
The rationale for implementing a "Minimum variant read count" filter twice in the pipeline (Steps 1 and 6) is as follows.
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