Biology · Biotechnology: Principles and Processes · NEET
DNA is cut at specific locations by restriction enzymes (restriction endonucleases), called "molecular scissors." Each enzyme reads a specific palindromic sequence of base pairs and cuts a little away from the centre of the palindrome, between the same two bases on both strands. This leaves short single-stranded overhangs called sticky ends. Memory hook: same enzyme = same sticky ends = the two DNA pieces can stick and join.
EcoRI reads the palindrome 5'-GAATTC-3' and cuts both strands between G and A, a little away from the centre. The offset cuts leave AATT single-stranded overhangs (sticky ends) that pair with complementary ends and are sealed by DNA ligase.
Your doubts, answered
Why do we cut DNA at a specific location and not randomly?
Because in genetic engineering we must join a chosen gene into a chosen vector in a controlled way. Restriction enzymes recognise a specific palindromic sequence and cut only there. This gives DNA fragments with known, matching ends. Random cutting would give unpredictable pieces that cannot be reliably rejoined.
What is a palindromic sequence in DNA?
It is a sequence of base pairs that reads the same on both strands when you read in the same 5' to 3' direction. Example: 5'-GAATTC-3' on the top strand has 3'-CTTAAG-5' below, and both read GAATTC in the 5' to 3' direction. This is like the word MALAYALAM, which reads the same forwards and backwards. Restriction enzymes recognise these palindromes.
Why does the same restriction enzyme have to cut both the gene and the vector?
When one enzyme (say EcoRI) cuts both the foreign DNA and the plasmid vector, both fragments get the SAME sticky ends. These complementary overhangs pair by hydrogen bonds, and DNA ligase seals them. If you used different enzymes, the sticky ends would not match and the pieces would not join.
Does a restriction endonuclease cut one strand or both strands?
It cuts BOTH strands of the DNA double helix. It cuts the sugar-phosphate backbone at a specific point on each strand. A common NEET trap says it cuts 'only one of the two strands' - that statement is wrong.
How are sticky ends formed?
The enzyme cuts a little away from the centre of the palindrome (not exactly at the centre), but between the same two bases on opposite strands. Because the two cuts are offset, short single-stranded stretches are left hanging. These overhangs are the sticky ends, named so because they form hydrogen bonds with complementary counterparts.
What is the difference between exonuclease and endonuclease here?
Both are nucleases. Exonucleases remove nucleotides from the ENDS of DNA. Endonucleases (which include restriction enzymes) make cuts at specific positions WITHIN the DNA. So the enzyme that cuts DNA at an internal specific site is an endonuclease, not an exonuclease.
⚠️ The NEET trap ✗ A restriction endonuclease binds DNA at a specific site and cuts only one of the two strands, exactly at the centre of the palindrome. ✓ A restriction endonuclease cuts BOTH strands of the double helix, cutting the sugar-phosphate backbone a little AWAY from the centre of the palindrome (between the same two bases on opposite strands), which leaves sticky ends. 🧠 Two words to remember: BOTH strands, and a little AWAY from centre. NTA repeatedly flips these into 'one strand' or 'exactly at the centre' to make a wrong option.
Real NEET questions
2022
In the following palindromic base sequence of DNA, which one can be cut easily by a particular restriction enzyme?
A · 5' GATACT 3' ; 3' CTATGA 5'
B · 5' GAATTC 3' ; 3' CTTAAG 5' ✓
C · 5' CTCAGT 3' ; 3' GAGTCA 5'
D · 5' GTATTC 3' ; 3' CATAAG 5'
Solution: Restriction enzymes recognise a true palindrome - a sequence that reads the same on both strands in the 5' to 3' direction. Only 5'-GAATTC-3' / 3'-CTTAAG-5' is a genuine palindrome (the EcoRI site); the others do not read the same on both strands, so they are not recognised.
2022
Statement I: Restriction endonucleases recognise specific sequences to cut DNA known as palindromic nucleotide sequences. Statement II: Restriction endonucleases cut the DNA strand a little away from the centre of the palindromic site. Choose the correct answer.
A · Both Statement I and Statement II are correct ✓
B · Both Statement I and Statement II are incorrect
C · Statement I is correct but Statement II is incorrect
D · Statement I is incorrect but Statement II is correct
Solution: Both statements are correct. Restriction endonucleases recognise a specific palindromic sequence (Statement I) and cut a little away from the centre of that palindrome, which produces the single-stranded sticky ends (Statement II).
2016
Which of the following restriction enzymes produces blunt ends?
A · Sal I
B · Eco RV ✓
C · Xho I
D · Hind III
Solution: Enzymes that cut both strands exactly at the centre of the palindrome leave flush (blunt) ends, while cutting away from the centre leaves sticky ends. EcoRV cuts at the centre and so produces blunt ends; Sal I, Xho I and Hind III produce sticky ends.
Solved Biotechnology: Principles and Processes NEET PYQs
Try the real previous-year questions from this chapter — each with the answer and a full solution.
They are called molecular scissors because they cut DNA at specific recognition sequences. The name 'restriction' comes from their original role of restricting the growth of bacteriophages in bacteria like E. coli.
What is the recognition sequence of EcoRI?
EcoRI recognises the palindromic hexamer 5'-GAATTC-3' with its complement 3'-CTTAAG-5'. It cuts between G and A on each strand, leaving AATT sticky ends.
Which enzyme joins the cut DNA fragments?
DNA ligase joins the fragments. After a restriction enzyme cuts the gene and the vector with matching sticky ends, DNA ligase seals the sugar-phosphate backbone to make recombinant DNA.
What is the difference between sticky ends and blunt ends?
Sticky ends have short single-stranded overhangs (enzyme cuts away from the centre), so they easily pair and join. Blunt ends are flush with no overhang (enzyme cuts exactly at the centre) and are harder to join.
Is a restriction endonuclease an exonuclease?
No. It is an endonuclease, which cuts within the DNA at a specific internal site. Exonucleases remove nucleotides from the ends of DNA - a different action.
