Last updated: 2017-12-13
Code version: 7509725
All processed data are stored as expressionSet objects in data/eset. Below shows how to extract information from an expressionSet object.
\(~\)
library(knitr)
library(Biobase)\(~\)
To combine all expressionSet objects in the folder,
fname <- Sys.glob("../data/eset/*.rds")
eset <- Reduce(combine, Map(readRDS, fname))\(~\)
To view the sample metadata labels,
kable(varMetadata(phenoData(eset)))| labelDescription | |
|---|---|
| experiment | ID of C1 chip (i.e. processing date in YYYYMMDD) |
| well | Well of C1 chip (96 total, rows A-H, cols 1-12) |
| cell_number | The number of cells observed in the well via microscopy |
| concentration | The cDNA concentration of the well prior to library prep |
| ERCC | The dilution factor of the ERCC spike-ins |
| individual.1 | Individual # 1 included on this C1 chip |
| individual.2 | Individual # 2 included on this C1 chip |
| image_individual | The chip label for the image files |
| image_label | The well label for the image files |
| raw | The number of raw reads |
| umi | The number of reads with a valid UMI |
| mapped | The number of reads with a valid UMI that mapped to a genome |
| unmapped | The number of reads with a valid UMI that did not map to a genome |
| reads_ercc | The number of reads that mapped to the ERCC spike-in transcripts |
| reads_hs | The number of reads that mapped to the H. sapiens genome |
| reads_egfp | The number of reads that mapped to the FUCCI EGFP transgene |
| reads_mcherry | The number of reads that mapped to the FUCCI mCherry transgene |
| molecules | The number of molecules (i.e. post UMI-deduplication) |
| mol_ercc | The number of molecules that mapped to the ERCC spike-in transcripts |
| mol_hs | The number of molecules that mapped to the H. sapiens genome |
| mol_egfp | The number of molecules that mapped to the FUCCI EGFP transgene |
| mol_mcherry | The number of molecules that mapped to the FUCCI mCherry transgene |
| detect_ercc | The number of ERCC genes with at least one molecule |
| detect_hs | The number of H. sapiens genes with at least one molecule |
| chip_id | verifyBamID: The predicted individual based on the sequencing data |
| chipmix | verifyBamID: chipmix is a metric for detecting sample swaps |
| freemix | verifyBamID: freemix is a measure of contamination. 0 == good & 0.5 == bad |
| snps | verifyBamID: The number of SNPs that passed thresholds for AF and missingness |
| reads | verifyBamID: The number of sequences that overlapped SNPs |
| avg_dp | verifyBamID: The average sequencing depth that covered a SNP |
| min_dp | verifyBamID: A minimun depth threshold for QC only (affects snps_w_min) |
| snps_w_min | verifyBamID: The number of SNPs that had the minimum depth (min_dp); QC only |
| valid_id | verifyBamID: Is the predicted individual 1 of the 2 added to the C1 chip? |
\(~\)
To view the feature (gene) metadata labels,
kable(varMetadata(featureData(eset)))| labelDescription | |
|---|---|
| chr | Chromosome |
| start | Most 5’ start position (GRCh37/hg19; 1-based; inclusive) |
| end | Most 3’ end position (GRCh37/hg19; 1-based; inclusive) |
| name | Gene name |
| strand | Strand (+ = positive/forward; - = negative/reverse) |
| source | Source of RNA |
\(~\)
To extract count data,
exprs(eset)There are 20,421 genes and 1,536 single cell samples in the raw data.
dim(exprs(eset))[1] 20421 1536\(~\)
To extract feature/gene information,
fData(eset)The features include FUCCI transgenes (EGFP and mCherry), ERCC spike-in controls, and endogenoeus genes (ENSG).
head(fData(eset)) chr start end name strand source
EGFP EGFP 1 714 EGFP + EGFP
ENSG00000000003 hsX 99883667 99894988 TSPAN6 - H. sapiens
ENSG00000000005 hsX 99839799 99854882 TNMD + H. sapiens
ENSG00000000419 hs20 49551404 49575092 DPM1 - H. sapiens
ENSG00000000457 hs1 169818772 169863408 SCYL3 - H. sapiens
ENSG00000000460 hs1 169631245 169823221 C1orf112 + H. sapiens\(~\)
To extract sample information,
pData(eset)The rows contain single cell samples. Row names indicate the experiment date (we had one C1 plate per day), and C1 well ID.
head(pData(eset)) experiment well cell_number concentration ERCC
20170905-A01 20170905 A01 1 1.7264044 50x dilution
20170905-A02 20170905 A02 1 1.4456926 50x dilution
20170905-A03 20170905 A03 1 1.8896170 50x dilution
20170905-A04 20170905 A04 1 0.4753723 50x dilution
20170905-A05 20170905 A05 1 0.5596827 50x dilution
20170905-A06 20170905 A06 1 2.1353518 50x dilution
individual.1 individual.2 image_individual image_label
20170905-A01 NA18855 NA18870 18870_18855 3
20170905-A02 NA18855 NA18870 18870_18855 2
20170905-A03 NA18855 NA18870 18870_18855 1
20170905-A04 NA18855 NA18870 18870_18855 49
20170905-A05 NA18855 NA18870 18870_18855 50
20170905-A06 NA18855 NA18870 18870_18855 51
raw umi mapped unmapped reads_ercc reads_hs
20170905-A01 2734844 1754078 1240443 513635 76433 1163764
20170905-A02 1910671 1254676 861713 392963 120589 740940
20170905-A03 2284182 1571727 1093848 477879 124186 968934
20170905-A04 920518 610742 382426 228316 116552 265873
20170905-A05 1260569 831378 494618 336760 117843 376703
20170905-A06 2501607 1733479 1258695 474784 99172 1158976
reads_egfp reads_mcherry molecules mol_ercc mol_hs mol_egfp
20170905-A01 246 0 113178 3122 110041 15
20170905-A02 182 2 59545 3143 56390 10
20170905-A03 727 1 74459 3307 71119 32
20170905-A04 1 0 29385 3110 26274 1
20170905-A05 71 1 42407 3059 39343 4
20170905-A06 546 1 94362 3120 91215 26
mol_mcherry detect_ercc detect_hs chip_id chipmix freemix
20170905-A01 0 39 8390 NA18870 0.12414 0.08025
20170905-A02 2 45 6057 NA18870 0.19067 0.10145
20170905-A03 1 40 6429 NA18855 0.21403 0.08767
20170905-A04 0 42 2746 NA18870 0.45097 0.08319
20170905-A05 1 44 3633 NA18870 0.44775 0.22013
20170905-A06 1 41 7508 NA18870 0.15767 0.11801
snps reads avg_dp min_dp snps_w_min valid_id
20170905-A01 311848 7959 0.03 1 3503 TRUE
20170905-A02 311848 3651 0.01 1 1802 TRUE
20170905-A03 311848 5059 0.02 1 2159 TRUE
20170905-A04 311848 815 0.00 1 591 TRUE
20170905-A05 311848 1209 0.00 1 780 TRUE
20170905-A06 311848 6722 0.02 1 2815 TRUER version 3.4.1 (2017-06-30)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Scientific Linux 7.2 (Nitrogen)
Matrix products: default
BLAS: /home/joycehsiao/miniconda3/envs/fucci-seq/lib/R/lib/libRblas.so
LAPACK: /home/joycehsiao/miniconda3/envs/fucci-seq/lib/R/lib/libRlapack.so
locale:
[1] LC_CTYPE=en_US.UTF-8 LC_NUMERIC=C
[3] LC_TIME=en_US.UTF-8 LC_COLLATE=en_US.UTF-8
[5] LC_MONETARY=en_US.UTF-8 LC_MESSAGES=en_US.UTF-8
[7] LC_PAPER=en_US.UTF-8 LC_NAME=C
[9] LC_ADDRESS=C LC_TELEPHONE=C
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
attached base packages:
[1] parallel stats graphics grDevices utils datasets methods
[8] base
other attached packages:
[1] Biobase_2.38.0 BiocGenerics_0.24.0 knitr_1.16
loaded via a namespace (and not attached):
[1] Rcpp_0.12.14 digest_0.6.12 rprojroot_1.2 backports_1.0.5
[5] git2r_0.19.0 magrittr_1.5 evaluate_0.10.1 highr_0.6
[9] stringi_1.1.2 rmarkdown_1.6 tools_3.4.1 stringr_1.2.0
[13] yaml_2.1.14 compiler_3.4.1 htmltools_0.3.6This R Markdown site was created with workflowr