normalization.Rmd

title: "single-cell RNA-Seq: Normalization"
author: "Laurent Modolo [laurent.modolo@ens-lyon.fr](mailto:laurent.modolo@ens-lyon.fr)"
date: "Wednesday 8 June 2022"
output:
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    theme: metropolis
  ioslides_presentation:
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classoption: aspectratio=169  

Introduction

Introduction

Program

1. Single-cell RNASeq data from 10X Sequencing (Friday 3 June 2022 - 14:00)
2. Normalization and spurious effects (Wednesday 8 June 2022 - 14:00)
3. Dimension reduction and data visualization (Monday 13 June 2022 - 15:00)
4. Clustering and annotation (Thursday 23 June 2022 - 14:00)
5. Pseudo-time and velocity inference (Thursday 30 June 2022 - 14:00)
6. Differential expression analysis (Friday 8 July 2022 - 14:00)

Introduction

Program

1. Single-cell RNASeq data from 10X Sequencing (Friday 3 June 2022 - 14:00)
2. Normalization and spurious effects (Wednesday 8 June 2022 - 14:00)

• Quality control
• Normalization
  • Variance stabilization
  • Depth normalization
  • The monotonicity of the normalization
• batch effects
• Heterogeneous data

3. Dimension reduction and data visualization (Monday 13 June 2022 - 15:00)
4. Clustering and annotation (Thursday 23 June 2022 - 14:00)
5. Pseudo-time and velocity inference (Thursday 30 June 2022 - 14:00)
6. Differential expression analysis (Friday 8 July 2022 - 14:00)

Quality control

Cell filtering

\begin{center} \begin{columns} \column{0.5\textwidth} \begin{center} \begin{tikzpicture} \fill (0.5,3.5) node {\bf \text{gene}_1} -- (0.5,2.5) node {\bf \text{gene}_2} -- (0.5,1.5) node {\bf \vdots} -- (0.5,0.5) node {\bf \text{gene}_n}; \fill (1.5,4.5) node {\bf{\text{cell}_1}} -- (1.5,3.5) node {mRNA} -- (1.5,2.5) node {mRNA} -- (1.5,1.5) node {\vdots} -- (1.5,0.5) node {mRNA}; \fill (2.5,4.5) node {\color{red}\bf{\text{0 cell}_2}} -- (2.5,3.5) node {\color{red}mRNA} -- (2.5,2.5) node {\color{red}mRNA} -- (2.5,1.5) node {\color{red}\vdots} -- (2.5,0.5) node {\color{red}mRNA}; \fill (3.5,4.5) node {\bf{\cdots}} -- (3.5,3.5) node {\cdots} -- (3.5,2.5) node {\cdots} -- (3.5,1.5) node {\ddots} -- (3.5,0.5) node {\cdots}; \fill (4.5,4.5) node {\bf{\text{cell}_c}} -- (4.5,3.5) node {mRNA} -- (4.5,2.5) node {mRNA} -- (4.5,1.5) node {\vdots} -- (4.5,0.5) node {mRNA}; \draw (1,0) grid (5,4); \end{tikzpicture} \end{center}

\column{0.5\textwidth}

{\large Some cells are not cells.}

\begin{itemize} \item matrix columns are defined by {\bf cell barcode sequences} \item {\bf cell barcode sequences identify droplet} in the 10X protocol \end{itemize}

\end{columns} \end{center}

Cell filtering

\begin{center} \begin{columns} \column{0.5\textwidth} \begin{tikzpicture} \fill (0.5,3.5) node {\bf \text{gene}_1} -- (0.5,2.5) node {\bf \text{gene}_2} -- (0.5,1.5) node {\bf \vdots} -- (0.5,0.5) node {\bf \text{gene}_n}; \fill (1.5,4.5) node {\bf \text{bc}_1} -- (1.5,3.5) node {mRNA} -- (1.5,2.5) node {mRNA} -- (1.5,1.5) node {\vdots} -- (1.5,0.5) node {mRNA}; \fill (2.5,4.5) node {\color{red}\bf \text{bc}_2} -- (2.5,3.5) node {\color{red}mRNA} -- (2.5,2.5) node {\color{red}mRNA} -- (2.5,1.5) node {\color{red}\vdots} -- (2.5,0.5) node {\color{red}mRNA}; \fill (3.5,4.5) node {\bf{\cdots}} -- (3.5,3.5) node {\cdots} -- (3.5,2.5) node {\cdots} -- (3.5,1.5) node {\ddots} -- (3.5,0.5) node {\cdots}; \fill (4.5,4.5) node {\bf \text{bc}_c} -- (4.5,3.5) node {mRNA} -- (4.5,2.5) node {mRNA} -- (4.5,1.5) node {\vdots} -- (4.5,0.5) node {mRNA}; \draw (1,0) grid (5,4); \end{tikzpicture}

\column{0.5\textwidth}