基因表达的表观调控
What turns a bee into a queen?
A typical eukaryotic gene structure
- TFs generally form a complex
- TF-binding just for seconds
- DNA organized in a compact form
- 表观修饰影响TF-binding and function
Epigenetics and epigenomics
Detect transcription factor - DNA interaction
- DNase I Footprinting Analysis
- Electrophoretic Mobility Shift Assay (Gel shift)
- Chromatin-Immunoprecipitation (ChIP) - Sequencing (ChIP-seq)
- Antibody-binding, sequencing, read peaks (Peak Calling)
- Cistrome Data Browser
- the detail info of every ChIP-seq experiment
Epigenetic marks:
- DNA methylation
- Histone modification
- H3K4me1/2/3, H3K27me1/2/3, H3K9me1/2/3
- Histone variant
- Nucleosome position and DNA looping
- higher-order chromatin structure
- …
the common nomenclature of histone modifications
- The name of the histone (e.g., H3)
- The single-letter amino acid abbreviation (e.g., K for Lysine) and the amino acid position in the protein
- The type of modification (Me: methyl, P: phosphate, Ac: acetyl, Ub: ubiquitin)
- The number of modifications (only Me is known to occur in more than one copy per residue. 1, 2 or 3 is mono-, di- or tri-methylation)
ATAC-seq: fast and sensitive profiling of open chromatin
- 可以与RNA-seq的data相结合，证明该代谢通路的chromatin被激活
最基本的测序思路，就是找出该区域的特异成分，bind到该region将其提取出来。
核酸是最大的数据库
蛋白质组学对研究技术的要求更高
相比于核酸，蛋白质只能做到「分离与检测」，核酸可以做到扩增放大、分离与检测、测序
蛋白质组学（proteomics）：以蛋白质组为研究对象，分析细胞内动态变化的蛋白质组成、表达水平和修饰状态
- 生物功能的主要载体
- 多样性及时空动态变化
液质联用（LC-MS/MS）：液相色谱+质谱
- 质谱：通过测定样品离子的质荷比（m/z）来进行成分和结构分析的方法
- 离子源⇒质量分析器⇒离子检测器
- MS/MS：串联质谱（Tandem MS），一级和二级质谱
- DDA （Data-dependent acquisition）vs DIA （Data-independent acquisition），DIA不管多少丰度全打出来，DDA可能会遗漏低丰度肽段
蛋白质组学技术应用
- 蛋白质组学检测系统组成
- 质谱基本原理
- 谱图解析原理
- 蛋白质组装和定量
CRISPR screening
Most targets were ignored in the pooled screens
ICRAFT: a systematic pipeline for gene target identification
Spatial sequencing, spatial tech benchmarking
spatial benchmarking of state-of-the-art sequencing methods