Question

A recombinant DNA is formed when sticky ends of vector DNA and foreign DNA Join. Explain how the sticky ends are formed and get jointed.

Hint:

Key Requirement: Use the {SAME} enzyme for both DNA sources.
If you use different enzymes, the sticky ends won't "match," and the DNA won't join.

Answer 1

Step 1: Understanding the Concept:
Genetic engineering requires "molecular scissors" (restriction enzymes) and "molecular glue" (DNA ligase).
Step 2: Detailed Explanation:
1. The Palindrome: Restriction enzymes recognize a "Palindromic Nucleotide Sequence"—a sequence that reads the same on both strands in the $5' \rightarrow 3'$ direction (e.g., GAATTC).
2. The Staggered Cut: Most restriction enzymes (like EcoRI) do not cut exactly in the middle of the palindrome. They cut the two strands slightly apart from each other.
3. Sticky End Formation: This results in short, single-stranded "tails" of DNA hanging off the ends. Because these tails have exposed bases, they are "sticky" (ready to base-pair with a complementary sequence).
4. The Joining Process:
- If both the source DNA and the vector DNA are cut with the same enzyme, their sticky ends will be perfectly complementary.
- They naturally find each other and form weak Hydrogen bonds.
- The enzyme DNA Ligase then makes the bond permanent by forming a covalent phosphodiester bond between the sugar and phosphate of the adjacent nucleotides.
Step 3: Final Answer:
Sticky ends are staggered cuts made by restriction endonucleases that facilitate the precise annealing of DNA fragments, which are then covalently sealed by DNA ligase.