bcbioSinglecell: Import and analyzebcbio single-cell RNA-seq data
Michael J. Steinbaugh, [email protected]
July 26, 2017
http://bioinformatics.sph.harvard.edu/bcbioSinglecell
Cell population-level analysis facilitates cell typeand differentiation cascade profiling
Stegle et al., Nat Rev Genet 16:133-145 (2015)
How to get here?Barcode filteringQuality control
§ Native HPC support for single-cell RNA-seq analysis.§ Uses the umis package internally to process barcodes.§ Quantifies counts using either rapmap or kallisto. § Outputs transcript-level counts in MartrixMarket format.§ bcbioSinglecell: R package interface to scRNA-seq data.
• Automates sample import into R as a SummarizedExperiment.• Saves counts as sparse matrix, using Matrix package.• Provides a number of quality control functions and RMarkdown
templates for clustering workflows (e.g. Seurat, monocle).• Data integration between Python and R!
https://bcbio-nextgen.readthedocs.io/http://bioinformatics.sph.harvard.edu/bcbioSinglecell/
Barcode samples
Sequence
FASTQ
Quasi-map reads
Process barcodes
inDropDrop-seqSureCell
Illumina
bcl2fastq
umis
rapmap, kallisto
scaterQuality control
Clustering
Differential expression
Trajectorymonocle
SCDE
Seurat
10X ChromiumCell Ranger
Load into R
bcbio
bcbioSinglecell
bcbioSCDataSet S4 class
gene-level counts dgCMatrix
Gene annotable
tx-level counts dgCMatrix
slotNames()
assays
metadata
callers
colData
NAMES
elementMetadata
bcbio run settings SimpleList
hdf5?
cellular barcode metrics
Ensembl gene identifiers
sparse = lower memory overhead
SummarizedExperiment
QC metrics and barcode filteringReads per barcode Cell counts
Gene detection Mitochondrial abundance
bcbioSCSubset S4 class
SummarizedExperiment
• min UMIs• min/max genes• max mito ratio• min novelty
seuratSCESet CellDataSet
bcbioSCDataSet
filter()
as()
bcbioSCSubset
scater Seurat monocle
Harvard Chan Bioinformatics Core
Mary Piper
John Hutchinson
Lorena Pantano
Brad Chapman
Peter Kraft
Rory Kirchner
Meeta Mistry Radhika Khetani
Shannan Ho Sui
Victor BarreraMichael Steinbaugh
