How to avoid Ns in small RNA reads or short reads after running bcl2fastq2?
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Small RNA reads are sometimes masked with Ns after FASTQ generation, resulting in the reads being 35 Ns instead of the actual basecalls. N-masking occurs when the read length is below the default minimum trimmed read length (ie, 35) and the adapter masking length (ie, 22) after adapter trimming.
To avoid N-masking in the reads, add the following to the bcl2fastq2 command line:
--minimum-trimmed-read-length 5
--mask-short-adapter-reads 5
More information about these options can be found in the bcl2fastq2 Conversion Software v2.20 User Guide.
For any feedback or questions regarding this article (Illumina Knowledge Article #3147), contact Illumina Technical Support .