Q.27 Which one of the following techniques is used to monitor RNA transcripts, both temporally
and spatially?
(A) Northern blotting
(B) In situ hybridization
(C) Southern blotting
(D) Western blotting
Answer: (B) In situ hybridization
In situ hybridization (ISH) detects specific RNA transcripts directly within intact cells or tissue sections, revealing both their spatial location (e.g., in specific cell types or organelles) and temporal expression patterns (e.g., during development or under stimuli).
Option Analysis
Northern blotting separates extracted RNA by size via gel electrophoresis, transfers it to a membrane, and hybridizes with labeled probes to quantify transcript levels and sizes, but it destroys spatial context from tissues.
Southern blotting targets DNA fragments after restriction digestion and electrophoresis for genetic analysis, not RNA transcripts.
Western blotting detects proteins using antibodies after SDS-PAGE separation, irrelevant to RNA monitoring.
Technique Overview
In situ hybridization uses labeled nucleic acid probes (DNA or RNA) complementary to target RNA sequences, applied to fixed tissues or cells; binding is visualized via fluorescence, chromogenic, or radioactive signals, preserving tissue architecture for spatial mapping.
In situ hybridization stands out as the premier technique to monitor RNA transcripts both temporally and spatially in molecular biology, enabling precise visualization of gene expression patterns within tissues.
Why In Situ Hybridization Excels
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Probes hybridize directly to RNA in fixed cells or sections, showing location (spatial) like neuronal expression in brain slices.
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Time-course experiments track changes (temporal), such as developmental upregulation.
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Variants like FISH (fluorescence) offer multiplexing for multiple transcripts.
Blotting Techniques Comparison
| Technique | Target | Spatial Resolution | Temporal Monitoring | Key Use |
|---|---|---|---|---|
| Northern Blotting | RNA | No (extracted) | Indirect via time points | Transcript size/quantity |
| In Situ Hybridization | RNA | Yes (in tissues) | Yes (dynamic imaging) | Localization patterns |
| Southern Blotting | DNA | No | No | DNA sequences |
| Western Blotting | Protein | No | Indirect | Protein levels |
Applications in Research
Researchers apply in situ hybridization to study embryogenesis, cancer microenvironments, and pathogen infections, where understanding RNA transcripts both temporally and spatially drives breakthroughs in biotechnology.
