Shown size is kb) and below uilis (total associated restriction sites present al

Question

Shown size is kb) and below uilis (total associated restriction sites present along the linear Referring to the figure below, predict the restriction fragment simplified version the genome ag boxes polymorphisms (RET that would result complete digests. Shade in on the agarose gel at the bottom of the page to the fragment present. The various shaded triangles indicate actual used the digest (in the the enzyme cut sites (in the diagram below) gel diagram at the bottom the page). filled in the answers for the first 2 anes in each gel as examples IN THE of FROM THE DIGESTS BELOW BANDS THAT RESULT Hind III cut site v- Dde I cut site v- Rsa cut site Taq I cut site 11 kb o kb 2. How would your answer differ if the genome of 1. This is for the linear genomeRFLP digest. Bacillus termaguilis (total size is 11 kb) was CIRCULAR? Indicate your answers below as you did for the linear genome above. A B C D E F G H

Explanation / Answer

We will see one by one restriction digestion of different enzyme and their cleaved product.

1 HindIII-

HindIII has only one site so when linear DNA is cleaved by HindIII we will two fragments one of the 4kb and another one of 7 kb.

In the case of circular DNA, we only get one fragment of 11kb as there is only one site for HindIII and upon digestion Circular DNA will be linearized.

DdeI - DdeI has three sites in the given DNA. So upon digestion of linear DNA, we will get four fragments of size -two fragments of 3kb, one of the 4kb and one of the 1kb.

upon digestion of circular DNA, we will get three fragments of size -one fragment of 3kb, two fragments of the 4kb

Rsa -

Rsa has four sites in the given DNA. So upon digestion of linear DNA, we will get five fragments of size - three fragments of two kb and one fragment each of 4kb, and 1kb.

upon digestion of circular DNA, we will get four fragments of size -two fragments of 3kb, two fragments of the 4kb and one fragment each of 2kb and 1 kb

Taq I-

Taq I has three sites in the given DNA. So upon digestion of linear DNA, we will get four fragments of size - one fragment each of 5kb, 3kb, 2kb, and 1kb.

upon digestion of circular DNA, we will get three fragments of size 5kb, 4kb and 2kb.

Double Digestion with Rsa and TaqI

Rsa has three sites and TaqI has also three sites.

One site of Rsa is overlapping with TaqI So we will get following fragments upon digestion of linear DNA- four fragments of 1kb, 2 fragments of 2 kb and one fragment of 3 kb

Upon digestion of circular DNA we will get following fragments- three fragments of 1kb, 1 fragments of 2 kb and two fragments of 3 kb.

Double digestion with HindIII and DdeI

HindIII have one site and DdeI has 3 sites. So we will get following fragments upon digestion of linear DNA- three fragments of 3kb and two fragments of 1kb

Upon digestion of circular DNA, we will get following fragments- two fragments of 3kb and one fragment of four kb one fragment of 1kb.

Triple digestion with HindIII, Taq I and DdeI

HindIII have one site, TaqI has three and DdeI has 3 sites. So we will get following fragments upon digestion of linear DNA-two fragments of 3kb, three fragments of 1kb and one fragment of 2kb

Upon digestion of circular DNA, we will get following fragments- one fragment of 4kb, one fragment of 3kb, two fragments of 1kb and one fragment of 2kb

Digestion with all the enzymes

We will get following fragments upon digestion of linear DNA- 2 + 1 + 3

one fragment of 3kb, six fragments of 1kb and one fragment of 2kb

Upon digestion of circular DNA, we will get following fragments- two fragments of 3kb, five fragments of 1kb

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