LAB - Crossing Over in Sordaria
Background: Refer to figure 3 as you read this background. Sordaria is a haploid (N) fungus in
the class Ascomycota. Like all members of this class, its spores are held in pea pod shaped cases
called asci. The gene for spore color is located on one of its chromosomes. In this lab, we will be
working with two different strains of Sordaria: a wild (black) spore producing strain and a mutant
(tan) spore producing strain. If the hypha of the different haploid (N) strains grow together and
make contact, their haploid nuclei can fuse together forming a diploid (2N) nucleus. After they
fuse, the cell will begin meiosis. If no crossing over occurs during meiosis, then the asci that form
will contain spores in a 4:4 pattern. If crossing over occurs we can have either a 2:4:2 pattern or a
2:2:2:2 pattern. The further the gene is from the centromere of the chromosome the greater the
chance of crossing over. So by determining the percentage of crossovers, we can calculate the
distance of the gene for spore color from the centromere. If we studied more than one gene, we
could then start to map the positions of the genes on the chromosome. Visit this website to
preview the images of the Sordaria asci http://www.jdenuno.com/PDFfiles/Sordaria.pdf .
Problem: You define the problem
Hypothesis: Will you be able to accurately determine the distance from the centromere to
the gene for spore color.
Variables: Think about this one
Materials: Crossing agar, Culture dish with crossing agar, scalpel, Bunsen burner, tooth pick,
slide, cover slip, dropper and water.
Procedures:
Preparing the Culture Dish
1. Do not open your culture dish until directed.
2. On the top near the edge of the dish mark your dish with your initials, period and
date.
3. On the bottom of your culture dish mark “+” for wild and “t” for tan in the four
places indicated by the diagram below.
4. Obtain a parent dish marked “+” for wild and a dish marked “t” for tan.
5. At your work station, flame your scalpel and then open the top of the parent dish
and remove one small block of agar containing the fungi.
6. Quickly, replace the cover of the parent dish.
7. Place the agar block (culture side down) on your crossing dish in the position
indicated in figure 1.
8. Repeat this for the second block of agar.
9. Flame the scalpel and then transfer two blocks of the other strain.
10. When you are finished the dish should look like figure 1 below.
** Avoid contamination keep covers on dishes.
Figure 1
11. Return your crossing dishes to the incubator.
8-10 Days Later
1. Use figure 2, on the next page, to determine the most probable area to find your
assigned mature perithecia.
2. Use a toothpick to remove a few mature perithecia (*) from the crossing plates and
make a wet mount.
3. Gently, press the coverslip down using the eraser end of a pencil to crush the
perithecia and release the clusters of the asci.
4. On 100X, examine the slide and locate rosettes of asci containing ascospores of two
different colors. The wild ascospores appear dark and both of the mutants
appear light gray or clear.
Figure 2
5. On 400X, examine the ascospores and determine if crossing over has taken place.
a. If crossing over has not taken place, the ascospores will be arranged in a 4:4
ratio.
b. If crossing over has taken place the ascospores will be arranged in a 2:4:2 or a
2:2:2:2 ratio.
c. See figure 3 on the next page.
d. Fill-in Table 1 and 2.
6. Write an appropriate conclusion, include all three parts. Think about the most
obvious source of error.
Data Collection:
Table 1 (Individual Data)
Table 2 (Class Data)
Data Processing and Presentation
Use the appropriate statistical tool to determine if any data is significantly different.
Use the appropriate statistical tool to determine if there is a significant difference between the
class data and the class expected data.
Use the appropriate statistical tool to determine if there is a significant difference between the
individual data and the individual expected data.
Conclusion and Evaluation
Don’t forget to address your hypothesis (no null hypothesis).
Using the known value for the gene to centromere distance, calculate the percent error for both
your data and class average data.
Figure 3