Due Thur. Nov. 23
An alternative approach is Radiation Hybrid (RH) mapping. In one method, cell lines are available in which one of the individual human chromosomes has been transferred to mouse cells by fusing human and mouse cells. To map a gene on one of the human chromosomes, choose the mouse cell line that contains the desired human chromosome, and treat with ionizing radiation. Ionizing radiation causes double-stranded breaks in chromosomal DNA, randomly distributed across the genome. Treated cells are next fused with drug-resistant mouse cells, and hybrids selected on fresh media. In many cases, a mouse cell will incorporate one or more broken human chromosomal fragments into mouse chromosomes, while most human chromosomes are lost. Several hundred clonal cell lines are established, each with one or more human fragments of a human chromosome. Some examples of clones are illustrated below (only human fragments are shown):
To use these cell lines for mapping, PCR-based markers of any kind can be used to score for presence or absence of loci. The closer together two loci are on the chromosome, the more frequently one will see bands for both loci. In the example above, A and B are found on three different fragments, while both A and D are both found on only one fragment. We would conclude that A is closer to B than it is to D.
a) Would the map distances calculated by RH be the same as distances calculated by Mendelian genetics? Why or why not?
b) Mendelian genetics requires the presence of two different alleles at each locus. Is this also true of RH markers? Explain.
2) (3 points)
Many species of bacteria can not be grown in pure culture, making them very difficult to study. This means that it is often difficult to estimate the number of bacterial species, and their relative abundance, in natural settings such as soil. If we make some simplifying assumptions, we could use C0t analysis to get an idea of the number of species in a soil sample. These assumptions are
all bacterial genomes are about the same size, using E. coli as the standard.
all bacterial genomes are distinct from one another ie. they do not cross hybridize.
In an experiment designed to assess the ecological effects of heavy metal pollution on bacteria, bacterial populations were isolated from soil samples at sites with high levels of metal contamination, low levels of contamination, or from uncontaminated soil. DNA was isolated from each bacterial sample, and C0t analysis was done, using purified E. coli DNA as a reference standard. The results are shown below.
Briefly describe the differences between bacterial populations in uncontaminated, low metal and high metal soils.