An analysis of chromosomal DNA using the Southern hybridization technique does not use:

Options

A. PCR
B. Electrophoresis
C. Blotting
D. Autoradiography

Answer

### Step-by-Step Solution:

1. **Understanding Southern Hybridization**: Southern hybridization is a technique used to detect specific DNA sequences in DNA samples. It involves several steps including the isolation of DNA, digestion with restriction enzymes, and the transfer of DNA fragments to a membrane.

2. **Isolation of Genomic DNA**: The first step in Southern hybridization is the isolation of chromosomal DNA from the cells. This is essential to obtain a pure sample for further analysis.

3. **Digestion with Restriction Enzymes**: After isolating the genomic DNA, it is digested using restriction endonucleases. These enzymes cut the DNA into smaller fragments, which is necessary for the subsequent separation step.

4. **Electrophoresis**: The resulting DNA fragments are then separated using gel electrophoresis. This process allows for the visualization of the DNA fragments based on their size.

5. **Blotting**: After electrophoresis, the DNA fragments are transferred to a nitrocellulose membrane. This process is known as blotting and is crucial for the hybridization step.

6. **Fixation and Blocking**: The DNA on the membrane is fixed, often by heating, and blocking agents are used to prevent non-specific binding during the hybridization process.

7. **Hybridization with Probes**: In Southern hybridization, a labeled probe (which is a single-stranded DNA) is introduced to the membrane. The probe will hybridize with the complementary DNA sequences present on the membrane.

8. **Autoradiography**: Finally, autoradiography is used to visualize the hybridized DNA. This technique involves exposing the membrane to X-ray film, which detects the radio-labeled probes.

9. **Identifying the Technique Not Used**: Among the options provided (PCR, electrophoresis, blotting, autoradiography), PCR (Polymerase Chain Reaction) is not used in Southern hybridization. PCR is a technique used for amplifying DNA, not for analyzing it after it has been isolated and digested.

### Final Answer:
The analysis of chromosomal DNA using the Southern hybridization technique does not use **PCR**.

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