BSCI330 EXAM 2 STUDY GUIDE Nucleus Questions 1 The composition of chromatin and comparison of genome sizes very broadly for viruses prokaryotes and eukaryotes level discussed in lecture Genome sizes E coli 4000 kb 1 36 mm circular human 2 900 000kb 990mm linear 23 chromosomes haploid virus From the table on slide 17 of Lecture Cell Nucleus SV40 Simian virus 5 1 kb 0 00017 mm circular prokaryotes nucleoid supercoiling via topoisomerases eukaryotes nucleus histones scaffolding note topoisomerase DNA gyrase exists in eukaryotic DNA as well but its function is to relieve tension generated by the unwinding of the helix at the transcription fork It does not generate a supercoil like topoisomerase I does in prokaryotes 2 Histones are small proteins with H2A core histone H2B core histone H3 tightly conserved between species core histone H4 tightly conserved between species with many basic amino acids that tend to be charged The charges on histones blunt the charges from phosphate groups on DNA and hold them together wouldn t basic amino acids imply a negative charge he specifically said that basic would cause a positive charge this makes sense since DNA is negatively charged At a neutral pH the basic amino acids Lys Arg are positively charged while the acids Glu and Asp would be negatively charged I assume when he says the histones have a positive charge this means there are more Lys Arg residues than there are Glu Asp H1 Linker histone keeps in place the DNA that has wrapped around the nucleosome H5 is also a linker histone 3 The folding or levels of condensation of a eukaryotic chromosome a starting with DNA and through the architecture of a metaphase chromosome plus b the experimental basis behind the information at the level of details discussed in lecture and related reading 5 steps to the folding of DNA DNA Double helix DNA wound around histones Fiber of tightly packed nucleosomes form by compacting histones Can be arranged in Zig Zag model two start or arranged in Solenoid model one Looping of chromosomes fibers of nucleosomes being wrapping condensing Called looped domains which are 300 nm thick and are attached to the scaffold Actual chromosome formed from multiple tight loop domains start Levels of condensation 1 DNA is packaged as a supercoil into nucleosomes 100 angstrom diameter interphase chromatin fiber Involves an octamer of histone molecules two each of histone H2a H2b H3 and H4 2 Additional folding and or supercoiling of the 100 angstrom nucleosome fiber to produce a 300 angstrom chromatin fiber characteristic of meiotic and mitotic chromosomes Histone H1 is involved in the supercoiling of the 100 angstrom fiber to produce the 300 angstrom fiber 3 Nonhistone chromosomal proteins form a scaffold that is involved in condensing the 300 angstrom chromatin fiber into the tightly packed metaphase chromosomes Appears to involve the segregation of segments of DNA molecules present in eukaryotic chromosomes into independently supercoiled domains or loops Mechanism unknown Experiment basically word for word from 10 2 lecture Take cells that are not in mitosis nuclei is disrupted with detergents to allow the material to relax and releasable Then washed with solns of moderate conc of ions other way is to wash them with more dilute soln lower ionic strength and see what happens then examined by e microscopy noticed beads on a string structure string had dimensions of dsDNA How would a sample of cells obtained from an individual be examined for potential chromosomal 6 abnormalities using karyotype analysis details of the assays and the reasons behind each of step Staining cells in mitosis using light microscopy First step in process is to take a growing population of cells Incubate the cells with colchicine up to 20 hours if they incubate longer they may not recover and perform apoptosis to allow cells to get to mitosis and get stuck so no matter what phase cells are in they will reach mitosis Cells will then accumulate in mitosis and begin to condense Cannot examine chromosomes until they condense and they don t begin to condense until they are in mitosis Mitosis is about 20 min process so are here for a very small amount of time by treating the cells with colchicine and accumulating them in mitosis you are increasing the chance of seeing stained chromosomes In mitosis phase you take the cells and shatter them before you fix them in methanol acetic acid to prohibit movement Then treat with trypsin protease solution to digest some of the proteins on the chromosome to allow the staining with Giemsa gives striped pattern seen on chromosomes Once you stain you look at what you see What does this sentence mean lol Take the cells under a light microscope and you see the chromosomes have separated and if you have the right spread enough separation you take a photograph of that section and with scissors cut the individual chromosomes on the printed photograph Line up the cut out chromosomes to make a karyotype and based on its size bending pattern and chromosome location one can identify a chromosome and make determination if there are shortened chromosomes or not Karyotype analysis lets us detect chromosomal abnormalities Question How would you use the spectral karyotyping technique to determine the chromosomal abnormalities If parts of chromosomes are lost or switched between chromosomes these changes can be detected by changes in the banding patterns or by changes in the pattern of chromosome painting pg 203 Spectral karyotyping shows each pair of chromosomes with a distinct color so if there are chromosomal abnormalities i e translocations there would be two different colors on an individual chromosome Look for abnormalities by observing missing material additional material or translocations BSCI330 Fall 2013 Review Questions Transport Give examples of different forms of facilitated diffusion that allow the movements of molecules through Facilitated diffusion is passive diffusion The movement of molecules according to their concentration gradient and requires no energy a an aqueous channel Aquaporins water channel proteins through which water molecules are able to pass through the phospholipid bilayer faster than by passive diffusion They are a type of glycerol channel glycerol channel glycerol glycine urea long sugar chains pass through ion channels mediate the passage of ions through the plasma membrane Channels are not always open Passage is mediated by binding of a
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