Question

A circular plasmid of 10,000 base pairs (bp) is digested with two restriction enzymes,A and B, to produce a
3000 bp and a 2000 bp bands when visualised on an agarose gel. when digested with one enzyme at a
time,only one band is visible at 5000 bp. If the first site for enzyme A(A1) is present at the 100th base, the
order in which the remaining sites(A2,B1 and B2) are present is?

Answer 1

Answer on Question#49517 - Biology - Molecular Biology

A circular plasmid of 10,000 base pairs (bp) is digested with two restriction enzymes, A and B, to produce a 3000 bp and a 2000 bp bands when visualised on an agarose gel. When digested with one enzyme at a time, only one band is visible at 5000 bp. If the first site for enzyme A(A1) is present at the 100th base, the order in which the remaining sites(A2,B1 and B2) are present is?

Answer

When the plasmid of 10,000 bp digested with one enzyme, gives 5000 bp band, it means that this plasmid has two restriction sites for this enzyme. If there were only one site for this enzyme, a single band of 10,000 bp would be produced. These two restriction sites are located 5000 bp far from each other.

When digested with two enzymes, these 5000 bp fragments are digested by the second enzyme, each of this fragment is cut by two, giving 2000 and 3000 bp fragments. Thus, this plasmid contains two restriction sites for the second enzyme. But there are two variants of restriction site location.

Possible restriction positions are listed in table below:

In both cases, restriction with two enzymes give 2 2000 bp fragments and 2 3000 bp fragments, while restriction with one enzyme at a time gives 2 5000 bp fragments.

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