Analysis of MiSeq Illumina data, how calculate the PF reads??

[mruizm]

Hi everyone, i'm analysing MiSeq Illumina data and i dont know how the software calculates the Pass Filtered reads, i was thinking that the total reads Pass Filtered was de 100% and the percentage of index was the number of reads, i.e.:

Total Reads PF: 1150491 -> 100%
Reads per Index: 46,0886%
Nº of reads for that index: X

Is that correct, to calculate the number of reads for each index??

[kcchan]

I'm not sure I understand your question. Are you asking about the statistics generated in either Sequence Analysis Viewer or in MiSeq Reporter?

[mruizm]

Hi kcchan, i'm asking about the Sequence analysis viewer statistics. What i want to calculate is the exactly number of reads that passed the filter. So when see the numbers in SAV the percentage shown in the % Reads Identified (PF) is not the percentage that correspond to the total PF reads, i'm calculating seeing the number shown on the Total Reads column. Do you understand?

[mruizm]

All of this information is shown in the Indexing QC section in SAV, by the way

[kcchan]

I would encourage you to read the User Guide for SAV; it describes each of the metrics in detail.

From the document:

• Total Reads—The total number of reads for this lane.
• PF Reads—The total number of passing filter reads for this lane.
• % Reads Identified (PF)—The total fraction of passing filter reads assigned to an index.

The % identified is the fraction of the PF reads that have been identified by the index that was listed on your sample sheet. This number will always be less than the % PF reads as some index reads have too many mismatches to be identified properly. Any PhiX control that was added in would also account for this difference.