Restriction analysis: Which enzymes recognize specific 4–8 base pair DNA sequences and cleave the phosphodiester backbone at or near those sites?

Introduction / Context:
Restriction endonucleases are sequence-specific DNA-cutting enzymes that launched the era of recombinant DNA. By cleaving at defined recognition sites (often palindromic), they generate sticky or blunt ends that enable precise cloning, mapping, and diagnostics. Their predictable activity is fundamental to constructing recombinant plasmids and analyzing genomes.

Given Data / Assumptions:

• Recognition sites are typically 4–8 base pairs, often palindromic in Type II enzymes.
• Cleavage can produce 5' overhangs, 3' overhangs, or blunt ends.
• Buffers, salt, and temperature affect activity and fidelity.

Concept / Approach:
The correct answer must specify enzymes that both recognize a specific DNA sequence and cleave the backbone accordingly. Restriction endonucleases do exactly this. Other DNA-associated enzymes unwind, ligate, supercoil, or synthesize DNA but do not perform site-specific restriction cutting for cloning purposes.

Step-by-Step Solution:

Verification / Alternative check:
Gel electrophoresis of digested DNA shows predictable fragment sizes that match theoretical maps, confirming site-specific cleavage by restriction endonucleases.

Why Other Options Are Wrong:

• DNA ligase: joins, does not cut.
• Helicases: unwind duplex DNA; no sequence-specific cutting.
• DNA gyrase: introduces negative supercoils in bacteria; not a cutter for cloning.
• Reverse transcriptase: synthesizes DNA from RNA templates.

Common Pitfalls:
Using the wrong buffer or methylation state can block restriction cutting; always verify dam/dcm sensitivity and star activity conditions.

Final Answer:
Restriction endonucleases