Question

Arrange the steps in southern blotting in correct order :
A. Transfer of DNA to nitrocellulose membrane
B. Detection of hybridized probe
C. Restriction digestion of DNA
D. Hybridization with labelled probe
E. Gel electrophoresis
Choose the correct answer from the options given below :

• A. \(A, C, D, E, B\)
• B. \(E, C, A, B, D\)
• C. \(C, E, A, D, B\)
• D. \(B, C, D, E, A\)

Hint:

Remember the sequence: \[ \text{Digest} \rightarrow \text{Separate} \rightarrow \text{Transfer} \rightarrow \text{Hybridize} \rightarrow \text{Detect} \]
• Southern blot \(\rightarrow\) DNA
• Northern blot \(\rightarrow\) RNA
• Western blot \(\rightarrow\) Protein

Answer 1

The Correct Option is C

Concept: Southern blotting is a molecular biology technique used for detection of specific DNA sequences. The technique involves:
• DNA fragmentation
• Separation by electrophoresis
• Transfer to membrane
• Hybridization with labelled probe
• Detection of target DNA It was developed by: \[ \text{Edwin Southern} \]

Step 1: Restriction digestion of DNA. Initially: \[ \text{DNA is cut into fragments using restriction enzymes} \] Thus, the first step is: \[ \boxed{C} \]

Step 2: Gel electrophoresis. The DNA fragments are then separated according to size using agarose gel electrophoresis. Thus: \[ \boxed{E} \]

Step 3: Transfer of DNA to membrane. Separated DNA fragments are transferred from the gel onto a nitrocellulose or nylon membrane. This process is called: \[ \text{Blotting} \] Thus: \[ \boxed{A} \]

Step 4: Hybridization with labelled probe. A labelled DNA probe complementary to the target sequence is added. The probe hybridizes with matching DNA fragments. Thus: \[ \boxed{D} \]

Step 5: Detection of hybridized probe. Finally: \[ \text{Hybridized probe is detected} \] using autoradiography, fluorescence, or chemiluminescence. Thus: \[ \boxed{B} \] Therefore, the correct sequence is: \[ \boxed{C \rightarrow E \rightarrow A \rightarrow D \rightarrow B} \] Hence, the correct answer is: \[ \boxed{(C)\ C, E, A, D, B} \]