NanoString Quality Control Dashboard

NanoString nCounter data are gene expression assays where there is no need for the use of enzymes or amplification protocols and work with fluorescent barcodes (Geiss et al. (2018) ). Each barcode is assigned a messenger-RNA/micro-RNA (mRNA/miRNA) which after bonding with its target can be counted. As a result each count of a specific barcode represents the presence of its target mRNA/miRNA. 'NACHO' (NAnoString quality Control dasHbOard) is able to analyse the exported NanoString nCounter data and facilitates the user in performing a quality control. 'NACHO' does this by visualising quality control metrics, expression of control genes, principal components and sample specific size factors in an interactive web application.

NAnoString quality Control dasHbOard

Installation

# Install the release from CRAN:
install.packages("NACHO")

# Or install the development version from R-universe:
install.packages(
  "NACHO",
  repos = c("https://mcanouil.r-universe.dev", "https://cloud.r-project.org")
)

# Or install the latest commit from GitHub with pak:
# install.packages("pak")
pak::pak("mcanouil/NACHO")

Quick start

The package ships six samples of the public study GSE178516, so this runs offline. load_rcc() reads the RCC files, computes the quality-control metrics and normalises the counts.

library(NACHO)

data_directory <- system.file("extdata", "GSE178516", package = "NACHO")
nacho <- load_rcc(
  data_directory = data_directory,
  ssheet_csv = file.path(data_directory, "samplesheet.csv"),
  id_colname = "IDFILE",
  instrument = "max",
  n_comp = 5
)
#> ℹ Reading 6 RCC files
#> ✔ Reading 6 RCC files [25ms]
#> 
#> ℹ Assembling 6 samples
#> ✔ Assembling 6 samples [18ms]
#> 
#> ℹ Computing quality-control metrics and normalising
#> ✔ Computing quality-control metrics and normalising [24ms]
#> 
nacho_qc(nacho)[, c("IDFILE", "status", "n_flags", "reason")]
#>                  IDFILE status n_flags reason
#> 1 GSM5393247_S02.RCC.gz   pass       0   <NA>
#> 2 GSM5393248_S13.RCC.gz   pass       0   <NA>
#> 3 GSM5393250_S15.RCC.gz   pass       0   <NA>
#> 4 GSM5393251_S16.RCC.gz   pass       0   <NA>
#> 5 GSM5393267_S46.RCC.gz   pass       0   <NA>
#> 6 GSM5393268_S47.RCC.gz   pass       0   <NA>

autoplot() draws any quality-control figure, render() writes a report with Quarto, and visualise() opens the app.

autoplot(nacho, type = "BD")

Box plot of the binding density of each cartridge, with the limits of the quality control drawn as dashed lines.

render(nacho, output_dir = tempdir())
visualise(nacho)

Overview

NACHO works with NanoString nCounter data. It reads the RCC files, flags the samples that fail quality control and normalises the counts.

load_rcc() reads the RCC files and a sample sheet, computes the quality-control metrics and normalises the counts.
visualise() opens a Shiny app to look at every metric and change the limits.
autoplot() draws any quality-control figure as a ggplot2 plot.
render() writes a quality-control report with Quarto, as an HTML or a PDF file.
normalise() runs the normalisation again with new settings.

The package website shows the app, the report and a full analysis.

Shiny application

Open the app on a nacho object with visualise().

visualise(GSE74821)

Short recording of the NACHO app on the example data. The summary strip changes when the field of view threshold moves to 95 and one sample is flagged. A selected sample is outlined in the QC metrics plots. The recording then shows the Samples, Normalisation and Batch pages, the Help menu, and the Export page with a report ready to download and the Reproduce in R card.

The articles “Explore quality control in the app” and “Analyse a GEO study” on the package website show each page of the app and a full analysis of GSE178516. The article “Moving from NACHO 2” on the same website maps the NACHO 2 code to NACHO 3.

Citing NACHO

Canouil, M., Bouland, G. A., Bonnefond, A., Froguel, P., ’t Hart, L. M., & Slieker, R. C. (2020). “NACHO: an R package for quality control of NanoString nCounter data”. Bioinformatics, 36(3), 970-971. doi: 10.1093/bioinformatics/btz647

@Article{Canouil2020,
  title = {{NACHO}: an {R} package for quality control of {NanoString} {nCounter} data},
  author = {Mickaël Canouil and Gerard A. Bouland and Amélie Bonnefond and Philippe Froguel and Leen M. {'t Hart} and Roderick C. Slieker},
  journal = {Bioinformatics},
  year = {2020},
  month = {feb},
  volume = {36},
  number = {3},
  pages = {970--971},
  issn = {1367-4803},
  doi = {10.1093/bioinformatics/btz647},
}

Getting help

If you encounter a clear bug, please file a minimal reproducible example on GitHub.
For questions and other discussion, please contact the package maintainer.