Question

The presence and distribution of specific mRNAs within a cell can be detected by

• A. Northern Blot analysis
• B. Rnase protection assay
• C. In situ hybridization
• D. Real-time PCR

Hint:

Whenever a question asks about the "location," "distribution," or "localization" of a molecule within a cell, look for techniques that involve microscopy or the term "in situ." Techniques that start with sample homogenization (like blotting or PCR) cannot provide this information.

Answer 1

The Correct Option is C

Step 1: Understanding the Concept:
The question asks for a technique that can determine not only if a specific mRNA is present but also where it is located ("distribution") within a cell or tissue. This requires a method that preserves the spatial integrity of the biological sample.
Step 2: Detailed Explanation:
Let's analyze the given techniques:
1. Northern Blot analysis: This method involves extracting total RNA from a population of cells, separating it by size on a gel, and then probing for a specific mRNA. It can tell you about the presence and quantity of an mRNA in the entire sample, but since the first step is to homogenize the cells, all information about the mRNA's original location is lost.
2. RNase protection assay (RPA): This is a sensitive solution-based method to detect and quantify specific RNAs. Like Northern blotting, it uses an RNA extract and provides no spatial information.
3. In situ hybridization (ISH): The term "in situ" is Latin for "in its original place." This technique uses a labeled probe (DNA or RNA) that is complementary to the target mRNA. The probe is applied to cells or tissues that have been fixed to preserve their structure. The probe hybridizes to the target mRNA, and its location is then visualized using microscopy. This directly reveals the spatial distribution of the mRNA within the cell or tissue. This perfectly matches the question's requirement.
4. Real-time PCR (or qRT-PCR): This technique is used to amplify and quantify the amount of a specific RNA (after converting it to cDNA) in a sample. It is highly quantitative but, like Northern blotting, requires RNA extraction, thus losing all spatial information.
Step 3: Final Answer:
In situ hybridization is the only technique among the options that allows for the detection and localization of specific mRNAs within their cellular context.