DNase I cuts DNA that is net protected by bound proteins but

DNase I cuts DNA that is net protected by bound proteins but is unable to cut DNA that is complexed with proteins. Human DMA is isolated, stripped of its nonhistone proteins, and mixed with DNase I. Samples are removed after 30 minutes. 1 hour, and 4 hours and run separately in gel electrophoresis. The resulting gel is stained with ethidium bromide, and the results are shown in the figure below. DNA fragment sizes in base pairs (bp) are estimated by the scale to the left of the gel. Examine the gel results and speculate why longer DNase I treatment produces different results. The rate of the enzymatic reaction under the experimental conditions is not sufficient to digest al DNA in 1 hour. The rate of the DNA-ethidium bromide interaction under the experimental conditions is not sufficient to stain all DNA in 1 hour The increase in size of 200 bp bancs is likely to result from denaturation of DNA under the experimental conditions. The decrease in size of 800 bp bands is likely to result from denaturation of DNA under the experimental conditions. Draw a conclusion about the organization of chromatin in the human genome from this gel. There are four types of nucleosomes that are located 200, 400, 600 and 800 bp apart. Nucleosomes are randomly distributed along DNA. Most nucleosomes are located 200 bp apart.

Solution

Part A. The human genome is very big and nucleosomes pack the genome into a compact structure. DNase I digestion is non-specific and cuts the genome at random size. That is reason after 30min digestion there are varied sizes of DNA on agarose gel. After 1 hr DNase I shreds the DNA into smaller pieces and by 3 hrs all the DNA which is not bound by nucleosomes are cut into pieces of 200 bps. So, the rate enzymatic reaction under the experimental conditions is not sufficient to digest all the DNA in an hour.

Part B: After 3 hrs of digestion you are able to see only DNA that are 200bps, which means they are protected by nucleosomes which are 200bps in length. After this time point, the DNase I cannot cut the DNA into smaller pieces.