Transcriptome: complete set of transcripts in a cell, for a specific developmental stage or physiological condition.
- interpreting the functional elements of the genome
- revealing the molecular constituents of cells and tissues
- understanding development and disease
RNA作为从DNA到protein的中间产物，能够起到上下信号的调控作用。protein已经是final product。
The aim of transcriptome study
- catalog all species of transcripts
- determine the structure of genes
Enhancer, promoter
CHIP-seq：研究TF结合的DNA序列
super-enhancer
3D genome
RNA-seq：转录组研究的主要技术
- Digital signal
- High sensitivity
- High Dynamic
mRNA肿瘤疫苗，至少解决了70%-80% hotspots
RNA递送问题，递送系统
RNA-seq data analysis: simple principle, but many methodological issues
- Read mapping
- Visualization
- Estimation: RPKM
- Discovery
Find differentially expressed genes: according to these types/classes, statistic analysis⇒
- different individuals/subtypes/situations/phases
- different genes in the genome/suspicious genes/selected genes
Then use:
- two classes: t-test, wilcoxon test
- multi-classes: ANOVA
除了p-value，还可以使用t-value等。
肿瘤微环境的免疫浸润细胞：使用RNA-seq研究免疫细胞的浸润
- 基因富集
- deconvolution
bulk RNA-seq
Single-cell RNA-seq
Droplet / microfluidics approaches
- amplification bias
- dropout rate ~ 90%
- background noise
- stochastic gene expression
- bias due to cell size, cell-cycle
Healthy vs Pathological tissues: 套路化
- understanding heterogeneous tissues
- identification and analysis of rare cell types
- changes in cellular composition
Branching
Seurat - single cell toolkit, scanpy
空间转录组学