Q.58 A student was asked to clone a DNA fragment, flanked by BamHI (GGATCC) sites into a vector, cut with Bglll (AGATCT).
The cloning was successful, as these two restriction enzymes are known to create compatible sites. Based on this, what will
be the restriction site configurations of these two enzymes (indicates the point of enzymatic cleavage).
1. G/GATCC and AGATC/T
2. G/GATCC and A/GATCT
3. GGATC/C and A/GATCT
4. GGATC/C and AGA/TCT
BamHI and BglII Compatible Restriction Sites in Cloning
Option 1: G/GATCC and AGATC/T correctly shows the cleavage patterns enabling successful ligation of BamHI-cut insert into BglII-cut vector.
Restriction Enzyme Basics
Type II enzymes BamHI (GGATCC) and BglII (AGATCT) generate compatible cohesive ends despite different recognition sequences.
Both produce 5′-GATC overhangs allowing sticky-end ligation, though hybrid sites resist recleavage by either enzyme.
This compatibility is exploited in directional cloning strategies.
Correct Cleavage Patterns
Option 1 matches standard configurations producing identical overhangs.
| Enzyme | Recognition | Cleavage Site | Overhang Generated |
|---|---|---|---|
| BamHI | GGATCC | G/GATCC | 5′-GATC |
| BglII | AGATCT | AGATC/T | 5′-GATC |
Ligation result: GGATCCT (BamHI-BglII junction) or AGATCTC—neither recreates original sites.
Option Analysis
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Option 1 (G/GATCC and AGATC/T): Correct—BamHI cuts after first G; BglII between 5th-6th bases, both yielding 5′-GATC sticky ends.
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Option 2 (G/GATCC and A/GATCT): Wrong—BglII slash position mismatches; produces incompatible TCTA overhang instead of GATC.
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Option 3 (GGATC/C and A/GATCT): Wrong—BamHI incorrectly shown cutting at 3′ end (GGATC/C), eliminating 5′-GATC overhang.
-
Option 4 (GGATC/C and AGA/TCT): Wrong—both enzymes misrepresented; no cohesive compatibility generated.
Why Compatibility Works
BamHI-cut: 5'-G GATCC-3'
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BglII-cut: 3'-CCTAG A -5'
Base-pairing of GATC overhangs enables ligation despite sequence differences—key concept for recombinant DNA technology in NEET/GATE biotech.
