Genetics: please show your work leading to the answer. Note that the answer is p

Question

Genetics: please show your work leading to the answer. Note that the answer is provided, I am just lost to how they got to the answer. Will give a thumbs up.

xtenued Caleulation A linear fragment of human DNA was isolated. The DNA molecule is digested with EcoRI and then, separately, with BamHI and HinDI. In addition, double digests are performed with these enzymes. The resulting fragments are run out on a gel. The pattern on the agarose gel is shown below. 10 kb . 10 kb 5 kb 4 kb 4 kb ??3kb 2 kb 1 kb B E H 5-3-1 E-3-1 2 How many EcoRI, BamHI and HinDIII sites are present in the DNA fragment? E= 2 B- 2 H-1 Draw a restriction map of the fragment. Indicate the restriction sites and the distances between the restriction sites. The gel was blotted to a membrane and probed with the 4.5kb EcoRI fragment. Which bands in the three double digests light up on the X-ray film? E/B- 2 and 2.5 B/H-6 and 4 E/H 4.5

Explanation / Answer

If a DNA has two restriction site, it will cut DNA at 2 sites and generate 3 fragments. Similarly when a restriction enzyme has one restriction site, it will cut the DNA at one place and will generate 2 fragments. Now, if we look on the agarose gel, EcoRI enzyme produces 3 bands on the gel, thus it will have 2 sites to cut the DNA. SImilarly, BamHI produces three fragments on the gel whereas, HindIII produces 2 frgments indicating bamHI has 2 sites and HindIII has one site to cut.

For analyzing restriction map, we need to see individually and also combined restriction cutting. For eg- E/B digestion three fragments has been generated, 4, 2.5 and 2. Thus when we mapped, E and B will ba mapped on DNA at a distance between 4Kb, 2.5Kb and 2Kb. On the other E alone will have one band with 4.5 Kb, one with 4Kb and last one 6.5 Kb.

If we probe the blot with 4.5Kb EcoRI fragment, it will includes E/B = 2 and 2.5 as both of these together will produce 4.5 Kb of single EcoRI digested fragment.

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