Question: I see the following error when demultiplexing my libraries: "The sample sheet supplied has a sample index collision". How can I continue to demultiplex my data?
Answer: This error suggests that the samplesheet supplied to mkfastq has a sample index ("barcode") collision. It can happen when the same sample index and lane were specified for multiple samples, or in certain cases where 10x Chromium i7 Multiplex Kit and i7 Multiplex Kit N samples were run on the same flowcell. It is a known issue that certain sample index combinations from these two different kits only differ by two bases-- meaning that it is possible for the sequencer to generate index reads with sequences that are one base away from multiple sample indices.
Please check your samplesheet to verify that you do not have duplicate lane-sample index pairs for multiple samples. If there are no duplicates, run
mkfastq with a
--barcode-mismatchs=0 argument. (The default parameter is
--barcode-mismatchs=1). This will make
bcl2fastq only accept reads that match the sample indices exactly. The small percentage of reads that are a single base away from multiple sample indices will be ignored.
You could also upgrade to most recent version of Cell Ranger which has a clearer message in case of sample index collision.