Q.42 A complete restriction digestion of a circular plasmid (5000bp) was carried out with
HindIII,BamHIandEcoRIindividually. Restriction digestion yielded following fragments.
Plasmid + HindIII→ 1200bp and3800bp
Plasmid + BamHI→5000bp
Plasmid + EcoRI→ 2500bp
The number of sites for EcoRI,BamHIandHindIIIpresent on this plasmid are
(A)EcoRI-2, BamHI-1, HindIII-2 (B)EcoRI-1, BamHI-1, HindIII-2
(C)EcoRI-3, BamHI-2, HindIII-1 (D)EcoRI-2, BamHI-2, HindIII-1

Decoding Plasmid Restriction Digestion: EcoRI, BamHI, and HindIII Sites Revealed

Plasmid restriction digestion analysis determines enzyme cut sites by fragment sizes from complete digests. For a 5000 bp circular plasmid digested individually with HindIII (1200 bp + 3800 bp), BamHI (5000 bp), and EcoRI (2500 bp), the correct site counts match option (A).

Core Principle

Circular plasmids yield n fragments from n restriction sites in complete digestion, as each cut linearizes and subsequent cuts divide the circle. Total fragment sizes always sum to plasmid length (5000 bp here). Single band equaling full size indicates no effective cuts.

HindIII Analysis

HindIII produces two fragments (1200 bp + 3800 bp = 5000 bp), confirming exactly 2 sites. One site would linearize to a single 5000 bp band; more than 2 would yield 3+ fragments.

BamHI Analysis

BamHI yields one 5000 bp fragment, indicating 0 or 1 site, but since no smaller pieces appear, effectively 1 site with no cut observed in this context—or standard interpretation as 1 absent/uncut site, but matching options confirm 1.

EcoRI Analysis

EcoRI shows one 2500 bp band, but for a 5000 bp circle, this implies two equal 2500 bp fragments from 2 sites directly opposite each other. One site would give 5000 bp linear; 3 sites would produce three fragments summing to 5000 bp, not one band.

Option Breakdown

• (A) EcoRI-2, BamHI-1, HindIII-2: Correct. Matches 2 EcoRI sites (two 2500 bp), 1 BamHI (uncut 5000 bp), 2 HindIII (1200+3800 bp).

• (B) EcoRI-1, BamHI-1, HindIII-2: Wrong. EcoRI-1 predicts single 5000 bp, not 2500 bp.

• (C) EcoRI-3, BamHI-2, HindIII-1: Wrong. EcoRI-3 needs 3 fragments; BamHI-2 needs 2; HindIII-1 needs single 5000 bp.

• (D) EcoRI-2, BamHI-2, HindIII-1: Wrong. BamHI-2 predicts 2 fragments; HindIII-1 predicts 5000 bp.