# DBT JRF 2020 — Question 37

Exam: DBT JRF 2020 · Section: A · Topic: Molecular Biology · Difficulty: Medium

## Question

Which of the following is a method of investigating the sequence specificity of DNbinding proteins in vitro?

## Options

- **A.** Southern hybridization
- **B.** DNA footprinting
- **C.** gene targeting
- **D.** polymerase chain reaction

## Answer

Option D

## Explanation

DNase I footprinting is used to identify the exact DNA sequence to which a protein binds. Method: (1) End-label the DNA fragment; (2) Incubate with the DNA-binding protein; (3) Treat with DNase I, which cleaves unprotected DNA at random positions; (4) Remove protein and run the DNA fragments on a gel. Where the protein was bound, DNase I cannot access — this region is 'protected' from cleavage and appears as a gap (footprint) in the ladder of bands. Comparing protein-bound vs. unbound lanes reveals the exact binding site sequence.

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