Question

Sequence of Southern Blotting is: A. Gel Electrophoresis, B. Hybridization with radioactivity labelled probe, C. Fragmentation with Restriction enzyme, D. Transfer to nitrocellulose filter, E. Autoradiophagy.

• A. A, E, B, C, D
• B. C, A, D, B, E
• C. A, B, C, E, D
• D. E, B, A, C, D

Hint:

Southern blotting sequence: Restriction digestion \(\rightarrow\) Gel electrophoresis \(\rightarrow\) Transfer to membrane \(\rightarrow\) Probe hybridization \(\rightarrow\) Autoradiography.

Answer 1

The Correct Option is B

Concept:
Southern blotting is a technique used to detect a specific DNA sequence in a DNA sample. It involves cutting DNA, separating fragments, transferring them to a membrane, hybridizing with a labelled probe, and detecting the signal.

Step 1: Fragmentation with restriction enzyme.
First, DNA is cut into smaller fragments using restriction enzymes. \[ \text{First step} = C \]

Step 2: Gel electrophoresis.
The DNA fragments are separated according to size by gel electrophoresis. \[ \text{Second step} = A \]

Step 3: Transfer to nitrocellulose filter.
The separated DNA fragments are transferred from the gel onto a nitrocellulose membrane or nylon membrane. \[ \text{Third step} = D \]

Step 4: Hybridization with labelled probe.
A radioactive or labelled probe is added. It binds to the complementary DNA sequence. \[ \text{Fourth step} = B \]

Step 5: Autoradiography.
Finally, autoradiography is used to detect the labelled DNA sequence. \[ \text{Fifth step} = E \] Therefore, the correct sequence is: \[ C, A, D, B, E \] \[ \therefore \text{Correct Answer is (B)} \]