Transcriptomics

trancriptomics is the study of all RNA molecules in a cell, tissure, or organism at a given time.

Transcriptome refers to:

the complete set of RNA molecules transcribed from the genome in a specific cell, tissue, or organism at a given time.

Concept	Description	Stability
Genome	All DNA sequences	Largely constant
Transcriptome	All transcribed RNA	Highly variable

Biological Significance

The transcriptome enables analysis of: - Gene expression patterns (which genes are active) - Expression levels (quantitative differences) - Cellular states (e.g., differentiation, stress response) - Regulatory processes (transcriptional and post-transcriptional control)

Experimental Approaches

• Historical:
  • Northern blot
  • RT-PCR
  • Microarrays
• Current Standard:
  • RNA-seq
  • Single-cell RNA-seq

W2kQ

All transcriptome methods are trying to answer: Which genes are expressed and how mcuh. But the dimentions are differnet: - Scale - resolution - quantitativeness - Bias

Detail Methods

Northern Blot

Brief Process:

1. Extract RNA
2. Separate by size (gel electrophoresis)
3. Transfer to membrane
4. Hybridize with labeled probe (complementary sequence)
5. Detect signal

Northern Blot Ability

• Able to detect presence of a specific RNA
• Able to determine RNA size (using RNA ladder)
• Able to detect splicing variants Able to measure approximate expression level based on signal strength
• Able to study same gene expression in different cell and tissues

Northern Blot limitations

• Low throughput (1 gene at a time)
• Require prior knowledge (probe design)
• Low sensitivity for rare transcripts
• Time consuming