Medical Archive

scRNA analysis

mRNA analysis pipeline of the NCI’s Genomic Data Commons for data reporcessing.

Quality Control

  1. The number of uniquely mapped read >15M
  2. Uniquely mapped reads >75% of the total aligned reads.

DEG Analysis

Significant Gene Selection

  • Default setting
    • Fold change: 2 / Normalized data (log): 4 / p-value: 0.05
    • Sample/control group : B/A both, C/A both, C/B both
  • Gene Category Chart
    • 각 GO관련 유전자 중 발현이 유의하게 차이 나는 유전자의 %와 수를 나타낸 그래프
  • Significant Chart
    • 선택한 비교조합에 따른 유의한 유전자(최대 30개)의 발현 값을 그룹 별로 확인 (p-value 순으로 표시)
Gene Category Chart / Significant Chart

Mutational Profile

Landscape of gene alterations in single cells (top 30). Tumor cells are sorted by radiosensitivity: RR vs. RS.
B.-S. Jang et al. / Radiotherapy and Oncology 142 (2020) 202–209

Mutational Signatures and Altered Pathway

B.-S. Jang et al. / Radiotherapy and Oncology 142 (2020) 202–209

Basic Plots

Scatter Plot

  • 대조군과 실험군의 발현양상을 확인할 수 있는 이미지
  • Setting: Sample/control group (e.g., B/A), fold threshold line (default: 2)

Volcano Plot

  • 반복 실험(N>=2)이 된 경우에만 분석 가능하다. Volcano Plot은 Scatter Plot의 기능과 거의 동일.

Venn Diagram

t-SNE

KEGG input

GSEA input

Selected Gene Plot

  • Select gene ID -> expression plot view

Radar Chart

Functional Annotation Analysis

DAVID

The Database for Annotation, Visualization, and Integrated Discovery

Start from extracting ‘DAVID input data’ from ExDEGA

https://david.ncifcrf.gov/

  • Start Analysis
  • Step 1: submit the gene list
    • 1. DAVID input file upload
    • 2. select official gene symbol (=identifier) (e.g., GENEBANK_ACCESSION)
    • 3. Gene list / Background
    • 4. convert list and submit to DAVID as a gene list
  • Step 2: analyze with one of DAVID tools
  • Step 3: functional annotation chart -> save the output file.

DAVID Graphic Analysis by ExDEGA GraphicPlus

Count of Gene
BubbleChart
P-Value
Radar

Clustering Heatmap Analysis

Clustering input

Producing ‘Clustering Heatmap Input.txt‘ file

  • Using GraphicPlus or MeV program
  • Type : Fold change / normalized data (Z-score) / average of normalized data (z-score)
  • Export Data Select : B/A, C/A, C/B

Hierarchical Clustering Heatmap by ExDEGA GraphicPlus

  • Upper dendrogram: sample cluster (normalized data with Z-score)
  • Leftside dendrogram: gene cluster

Dimensionality Reduction

PCA (Principal Component Analysis)

from <B-Cell Immunity Predicts ER-Positive Breast Cancer Prognosis>

UMAP

From Molecular Therapy Vol. 32 No 11 November 2024

String Network Analysis

  1. Select genes of interest (<100)
  2. Draw -> saved as .svg file

SVG file

Excel file

  • Node 1, node 2 and the interaction score between the two.

Correlation Analysis

HeatMap / PairGrid / Mix

Pathway Analysis

Using KEGG Mapper:

Gene Set Enrichment Analysis (GSEA)

Microarray or RNA-seq data -> significant gene set analysis

GSEA report files

  • ‘gsea_report_for_A_000’ : control group’s enriched gene set
  • ‘gsea_report_for_B_000’: experimental group’s enriched gene set

Protein-Protein Network Analysis

Cytoscape STRING tool lets us identify the protein-protein interaction based on the database.