bcbiosinglecell: import and analyze bcbiosingle-cell rna-seq...
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bcbioSinglecell: Import and analyzebcbio single-cell RNA-seq data
Michael J. Steinbaugh, [email protected]
July 26, 2017
http://bioinformatics.sph.harvard.edu/bcbioSinglecell
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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
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§ 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/
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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
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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
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QC metrics and barcode filteringReads per barcode Cell counts
Gene detection Mitochondrial abundance
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bcbioSCSubset S4 class
SummarizedExperiment
• min UMIs• min/max genes• max mito ratio• min novelty
seuratSCESet CellDataSet
bcbioSCDataSet
filter()
as()
bcbioSCSubset
scater Seurat monocle
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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