BIO 344 Quiz 3

1. DNA PACKAGING-Fill in the blanks for the following summary about DNA packaging.

Chromosomes are made up of __chromatin______, which is DNA+protein. _Histones______proteins make up most of the protein component. There are five of these proteins, a core complex containing two copies each of _H2a__, H2b__, _H3___, and _H4__, and one copy of _H1__ which holds several cores together. When associated with DNA, this protein-DNA complex is called a/an _nucleosome______and is the first level of compaction. The second level of compaction is called the 30nm_fiber_or solenoid______and contains 6 _nucleosome__s per turn. The next level of compaction involves the looping of the _solenoid______in and out, using a _scaffold______to attach to. Regions near the scaffold have ___higher ______levels of transcription while those in the loops have __lower______levels. At this point we are basically looking at euchromatin, or interphase chomosomes. Further condensation during metaphase allows us to actually visualize chromosomes in the light microscope.

2.EXPERIMENT

EM’s of the SV40 virus show that promoter regions are typically void of nucleosomes. This could explain how transcriptional activation at these sites can occur: no nucleosomes, no problem for access of RNA polymerase. However we do see DNA complexed with histone (i.e. nucleosomes) throughout the rest of the gene downstream. In class we discussed an experiment to determine what happens when RNA polymerase encounters nucleosomes during elongation. Answer the following questions about this experiment.

A. What are two possible ways that the polymerase can deal with the nucleosomes, i.e. the fundamental question of this experiment?

1, Slide on the DNA which is around it.

2. Cause their dissembly.

B. Briefly explain construction of their “template” or starting material and how it could be used

to get a result.

The template is a plasmid that a 5S rRNA gene with only one nucleosome placed downstream of a promoter. After transcription, the DNA protected by the nucleosome is recovered, radiolabeled and hybridized to the plasmid digested with several restriction enzymes (to see if the nucleosome moved elsewhere in the plasmid).

C. How does micrococcal nuclease work and why is it important to this experiment?

MN degrades all the plasmid DNA except what’s in a nucleosome.

D. Why is it essential to probe against restriction enzyme fragments of the plasmid?

To see where what part of the plasmid is in a nucleosome after transcription.

3.DNA SUPERCOILING

Write out the equation that describes linking number, defining each variable.

Lk= Tw + Wr

Lk = total # of times that strands cross

Wr= # of times that the double helix crosses itself

Wr= # of times that the strands cross, but excluding writhe

4.RECOMBINATION

Draw out the pathway of recBCD homologous recombination

For next test.