Import and analyze bcbio single-cell RNA-seq data.

Installation

This is an R package.

Bioconductor method

source("https://bioconductor.org/biocLite.R")
biocLite("ensembldb")
biocLite(
    "hbc/bcbioSingleCell",
    dependencies = c("Depends", "Imports", "Suggests")
)

Sample metadata examples

FASTQ files with samples multiplexed by index barcode

This is our current method for handling inDrop samples.

fileName description index sequence sampleName
170201_R1.fastq.gz run1 17 GGAGGTAA sample1
170201_R1.fastq.gz run1 18 CATAACTG sample2
170620_R1.fastq.gz run2 12 GCGTAAGA sample3
170620_R1.fastq.gz run2 13 CTATTAAG sample4
170620_R1.fastq.gz run2 14 AAGGCTAT sample5
170620_R1.fastq.gz run2 15 GAGCCTTA sample6
170620_R1.fastq.gz run2 16 TTATGCGA sample7

FASTQ files demultiplexed per sample

This is our current method for handling 10X and SureCell samples.

fileName description
sample1_170620_R1.fastq.gz sample1
sample2_170620_R1.fastq.gz sample2
sample3_170620_R1.fastq.gz sample3
sample4_170620_R1.fastq.gz sample4
sample5_170620_R1.fastq.gz sample5

Technical replicates

Use sampleNameAggregate to assign groupings for technical replicates:

fileName description sampleNameAggregate
wildtype_L001_R1.fastq.gz wildtype_L001 wildtype
wildtype_L002_R1.fastq.gz wildtype_L002 wildtype
wildtype_L003_R1.fastq.gz wildtype_L003 wildtype
wildtype_L004_R1.fastq.gz wildtype_L004 wildtype
mutant_L001_R1.fastq.gz mutant_L001 mutant
mutant_L002_R1.fastq.gz mutant_L002 mutant
mutant_L003_R1.fastq.gz mutant_L003 mutant
mutant_L004_R1.fastq.gz mutant_L004 mutant

Troubleshooting

Maximal number of DLLs reached

Error: package or namespace load failed for 'bcbioSingleCell' in dyn.load(file, DLLpath = DLLpath, ...):
  maximal number of DLLs reached...

Depending on your operating system, you may encounter this error about hitting the DLL limit in R. This issue is becoming more common as RNA-seq analysis packages grow increasingly complex. Luckily, we can configure R to increase the DLL limit. Append this line to your ~/.Renviron file:

R_MAX_NUM_DLLS=150

For more information on this issue, consult help("dyn.load") in the R documentation. The number of loaded DLLs in an R session can be obtained with getLoadedDLLs().