Question - Why do I have an error "Sequence and quality not separated by +" while running 10x Genomics Cloud Analysis?
Answer - The following error "Sequence and quality not separated by +" on Cloud Analysis indicates a possible issue with the input FASTQ files.
[error] Pipestance failed. Error log at: 0070c59a4b98-a055ce7af46d9f5e/SC_RNA_COUNTER_CS/SC_MULTI_CORE/MULTI_GEM_WELL_PROCESSOR/COUNT_GEM_WELL_PROCESSOR/_BASIC_SC_RNA_COUNTER/_MATRIX_COMPUTER/MAKE_SHARD/fork0/chnk2-z7e2a757b73/_errors Log message: Sequence and quality not separated by +: file: "/mnt/scratch/inputs/762d56b8a5a4c1ad5e1b2a63b1d/20f1d7b09721-26f812e71eeb/Sample_S1_L002_R2_001.fastq.gz", line: 930524 Pipestance failed, staying alive because --noexit given.
Cloud Analysis (and 10x Ranger pipelines) require the input FASTQ files to be complete and uncorrupted. All reads in a given FASTQ fileset are expected to follow the FASTQ format for R1/2/3 and I1/2 (if present).
If you encounter the error described in the article, you will need to verify the integrity of your FASTQ files using md5sum before and after downloading the files. It is expected to have the same number of entries in R1/2 and I1/2 for 10x datasets and performing a line count is a quick way to check whether the files are truncated or not.
Cloud Analysis is only available in the United States and Canada.