Alex Wilcox
October 22, 2014
Sordaria Lab Write Up Summary
1.
Introduction a.
Background information about phenomenon of increased cross-over frequency found in SFS compared to NFS: SFS receives more radiation than the NFS. SFS has a higher likelihood of drought and has higher temperatures than the NFS. Differences in environmental conditions have led to researchers desiring to investigate crossover frequencies between organisms in each of the slopes. In this experiment, they used Sordaria, which are seen as good model organisms since they have fast generation times, and it is easy to isolate the different kinds or strains of the fungi.
(pages 34 and 35 in lab notebook) b.
Need for baseline information about cross-over frequencies of the spore color gene from populations that have been grown under optimal lab growth conditions: There is information from the Evolution Canyon that there was a decent amount of differences in the cross-over rate between the North and South facing slopes. The cross-over frequencies were determined between spore color mutant and wild type strains of the
Sordaria. It will be necessary to see whether there is a difference between the cross-over frequencies in natural population settings and under lab conditions before one can compare these two slopes though. The lab conditions will represent the baseline information for the experiment.
Scientists desire to set up an experiment that allows for a control group in part of the overarching experiment. (pages 35 and 36 in lab notebook). c.
Background information about types of asci that can occur (4:4. 2:2:2:2,
2:4:2) and how each can arise: In the Sordaria, when the wild and mutant strains undergo sexual reproduction there can be different arrangements of the ascospores in the ascus. When the asci undergo meiosis followed by mitosis there is a result of 8 daughter ascospores. If there is no crossover or genetic recombination the arrangement that will occur after meiosis and mitosis has occurred is 4:4. If the right chromatid of either the wild or the mutant strain crosses over with the left chromatid of the other strain in prophase I of meiosis, than it will result in a 2:2:2:2 formation in the ascus. Finally, if the left chromatid of either the wild or mutant strain crosses over with the left chromatid of the opposite strain in prophase I of meiosis (or right with other right), the result will be a
2:4:2 formation in the ascus. (pages 48-51 in lab notebook) d.
Research Questions for the experiment: (page 37 in lab notebook) i: Does crossing-over occur between the wild color gene and the centromere? ii: What is the frequency of crossing-over between the wild color gene and the centromere in the Sordaria grown under normal laboratory conditions?

iii: What is the map distance that is calculated between the wild color gene and the centromere? e.
Brief overview of procedures you used and what you will be looking for. i: Crosses that were done: Cross-over between the wild type and tan type of Sordaria under room temperature after 2 weeks in a agar plate. We will be looking for the cross-over rates between the quadrants. (page 42 in lab notebook) ii: Cross-over frequencies that were examined (frequency of each cross-over type and overall crossover frequency): need to determine whether there is a higher frequency of 2:4:2 or 2:2:2:2 formations in the ascus after crossing-over has occurred, and then compare it to the frequency of when the Sordaria did not cross-over (4:4 formation in ascus). Finally we will need to compare the frequency of crossing-over under optimal lab conditions to that of the Evolution Canyon slopes. (pages 36 and 42 of lab notebook)
2.
Methods a.
Describe how the crosses were set up: We started off by drawing two straight lines across a mating agar plate to design four quadrants. We wrote down “Tan” and “Wild type” alternatively in the four different quadrants. Here, we made sure they were alternate so a “Tan” was diagonal and not across from another “Tan type” and the same for the
“Wild type” of the Sordaria. Next we made sure to disinfect and clean the work surface and the scalpel that we would use to cut squares of agar with. One lab member cut two squares of either the “Wild” or “Tan” from a fungal culture plate. She then placed them accordingly on a quadrant diagonal from each other with the fungal side down. Then the lab member repeated this process with the opposite type of Sordaria. Finally, we disinfected the scalpel that we had used in the experiment. (page 42 of lab notebook) b.
Determine how squashes were prepared: The squashes were prepared starting off with the use of an inoculating loop to scrape part of the perithecia from the agar plate. They then were placed onto the microscope slide in a drop of water. After the slide gets covered with a coverslip a group member pressed down gently with the eraser part of a pencil to remove the asci from the perithecia. Next in the experiment, the coverslip needed to be slid around to make sure that the asci got spread out on it and finally the coverslip could be put under a microscope to view the asci. Once the group discovered a group of asci under the 100X magnification, they could be viewed under the 400X magnification to look for the three different types of asci. Concluding this part of the experiment, each group member could count various types of asci on the coverslip. (page 56 of lab notebook). c.
Describe how cross-over frequencies and map distances were calculated:

The cross-over frequencies were calculated by dividing the total number of recombinant asci by the total number of asci counted. The map distances could then be calculated by dividing the cross-over frequency percentage by 2 since only one chromatid (or half the chromosome) of each type could cross-over with each other in the Sordaria.
3.
Results a.
Provide a table showing class results for: i: number of asci scored for each ascus type ii: total number of asci scored
Class Results
Total # of Asci # of Type A Asci
4:4
120
# of Type B Asci
2:4:2
109
# of Type C Asci
2:2:2:2
116 b.
Provide a table for cross-over frequencies showing: i: Crossover frequency for each cross-over type ii: Overall cross-over frequency
Cross-Over Frequency for Class
225
Frequency of Type B
Asci (2:4:2)
31.6%
Frequency of Type C
Asci (2:2:2:2)
33.6%
Frequency of Crossover
65.2% c. Written description of patterns observed that relate to the research questions i: Does crossing-over occur between the spore color gene and the centromere? Yes there was a significant amount of crossing-over that occurred between the spore color gene and the centromere. ii: What is the overall cross-over frequency between the spore color gene and the centromere in organisms grown under standard laboratory growth conditions? The overall cross-over frequency between the spore color gene and the centromere in laboratory conditions in my section was
65.2%. The overall cross-over frequency for Biology 110 course was iii: What is the calculated map distance between the spore color gene and the centromere? To calculate the map distance one must determine the percent cross-over and divide it by two. It must be divided by 2 because only one chromatid from each chromosome can cross-over during meiosis I in
Sordaria. The calculated map distance for my section was 32.6 mu away from centromere. The calculated map distance for the entire bio 110 course was
4.
Discussion a.
Claims and evidence (Statements of what you learned and the evidence from patterns you observed that supports those claims): I learned that

even in near perfect conditions for Sordaria, the fungal species will still have cross-over occur during mating. The fact that my section had a cross-over rate of 65.2% is pretty significant. I also learned about why there is more likely to be a 2:2:2:2 than a 2:4:2 combination of ascospores due to it being easier to cross-over this way due to the positioning of the homologous chromosomes in prophase I of meiosis. My results verified the fact that the 2:2:2:2 combinations was more likely. b.
Discussion of unexpected findings: I believed that it was very surprising that there was such a high rate of crossing-over under laboratory conditions. I expected there to be a significant lower rate of it due to the fact that it was nearly perfect conditions for the Sordaria. I also found it interesting that there was a higher likelihood that there would be a
2:2:2:2 combination than a 2:4:2 combination from crossing-over. c.
Implications of information learned for further experiments: I think it would be important to conduct an experiment on both the North and
South facing slope now that the baseline information for a control group in the experiment exists. It would be interesting to see how different the rate of cross-over is between the African-like slope (South), the
European-like slope (North), and the controlled laboratory setting.