lab meiosis

AP Biology Name_____________________ Meiosis Lab Determine the cross over frequency in the fungus Sordaria fimicola. Use the cross-over frequency to make a chromosome map Observe prepared slides of the results from a cross between wild type fungus (black spores) and a mutant (tan spores) Count Asci that have both black and tan spores. Calculate cross over frequency, x?, % error (individual and class) Background: Write a few (2-3) introductory paragraphs that provide background information about meiosis. Use the following questions to guide you. What type of cell is formed by meiosis? What is the purpose of those cells? What are the following? ASCUS, ASCOSPORES, PERITHECIUM Why are there 8 spores in an ascus? Summarize the key points of the Sordaria life cycle and crossing over. Use the background information provided below or found in your textbook and online. Crossing Over during Meiosis in Sordaria Sordaria fimicola is an ascomycete fungus that can be used to demonstrate the results of crossing over during meiosis. Sordaria is a haploid organism for most of its life cycle. It becomes diploid only when the fusion of the mycelia (very small filaments) of two different strains results in the fusion of the two different types of haploid nuclei to form a diploid nucleus. The diploid nucleus must then undergo meiosis to resume its haploid state. Meiosis, followed by mitosis, in Sordaria results in the formation of eight haploid ascospores contained within a sac called an ascus (plural, asci). Many asci are contained within a fruiting body called a perithecium. When ascospores are mature the ascus ruptures, releasing the ascospores. Each ascospore can develop into a new haploid fungus. Sordaria growing on a plate is shown in figure 1, Figure 2 shows a ruptured perithecium and acsi with spores. Figure 3 shows the lifecycle of Sordaria. Fig. 1:Sordaria growing on plate Figure 2: Ruptured perithecium, asci with spores Figure 3: Life cycle of Sordaria To observe crossing over in Sordaria, one must make hybrids between wild-type and mutant strains of Sordaria. Wild-type (+) Sordaria have black ascospores. One mutant strain has tan spores (tn). When mycelia of these two different strains come together and undergo meiosis, the asci that develop will contain four black ascospores and four tan ascospores. The arrangement of the spores directly reflects whether or not crossing over has occurred. In Figure 2, no crossing over has occurred. Figure 4 shows the results of crossing over between the centromere of the chromosome and the gene for ascospore color. Fig. 4: Formation of Non-crossover Asci Two homologous chromosomes line up at metaphase I of meiosis. The two chromatids of one chromosome each carry the gene for tan spore color (tn) and the two chromatids of the other chromosome carry the gene for wild-type spore color (+). The first meiotic division (MI) results in two cells each containing just one type of spore color gene (either tan or wild-type). Therefore, segregation of these genes has occurred at the first meiotic division (MI). The second meiotic division (MII) results in four cells, each with the haploid number of chromosomes (lN). A mitotic division simply duplicates these cells, resulting in 8 spores. They are arranged in the 4:4 pattern. Fig. 3: Formation of Crossover Asci In this example, crossing over has occurred in the region between the gene for spore color and the centromere. The homologous chromosomes separate during meiosis I. This time, the MI results in two cells, each containing both genes (1 tan, 1 wild-type); therefore, the genes for spore color have not yet segregated. Meiosis II (MII) results in segregation of the two types of genes for spore color. A mitotic division results in 8 spores arranged in the 2:2:2:2 or 2:4:2 pattern. Any one of these spore arrangements would indicate that crossing over has occurred between the gene for spore coat color and the centromere. Two strains of Sordaria (wild-type and tan mutant) have been inoculated on a plate of agar. Where the mycelia of the two strains meet (Figure 4), fruiting bodies called perithecia develop. Meiosis occurs within the perithecia during the formation of asci. A slide has been prepared of some perithecia (the black dots in figure 4). Fig. 4 Materials and Methods Obtain a prepared slide that show several “squished” perithecia that resulted from a fusion of black wild type and tan mutant haploid mycelia. Focus at 400x. Each perithecium contains many acsi; each ascus contains 8 ascospores. Your job is to count at least 50 hybrid asci, determining the #asci with non-crossover (4:4) and crossover (2:4:2 or 2:2:2:2) patterns within the asci. Enter your data in a table. (see below for example) Calculations for the tale: % Crossover = # Crossover asci divided by total #asci X 100 Map Distance = % Crossover asci divided by 2 (to account for the effects of mitosis – only half of the spores in each ascus are the result of meiosis) Table # Non-Crossover Asci (4:4) # Crossover Asci (2:4:2 or 2:2:2:2) Total # Asci % Asci showing crossover Gene to Centromere Distance (Calculated as % Asci showing crossover ÷ 2 (Centimorgans)) The frequency of crossing over appears to be governed largely by the distance between genes, or in this case, between the gene for spore coat color and the centromere. The probability of a crossover occurring between two particular genes on the same chromosome (linked genes) increases as the distance between those genes becomes larger. The frequency of crossover, therefore, appears to be directly proportional to the distance between genes. A map unit is an arbitrary unit of measure used to describe relative distances between linked genes. The number of map units between two genes or between a gene and the centromere is equal to the percentage of recombinants. Customary units cannot be used because we cannot directly visualize genes with the light microscope. However, due to the relationship between distance and crossover frequency, we may use the map unit. The frequency of crossing over appears to be governed largely by the distance between genes, or in this case, between the gene for spore coat color and the centromere. The probability of a crossover occurring between two particular genes on the same chromosome (linked genes) increases as the distance between those genes becomes larger. The frequency of crossover, therefore, appears to be directly proportional to the distance between genes. A map unit is an arbitrary unit of measure used to describe relative distances between linked genes. The number of map units between two genes or between a gene and the centromere is equal to the percentage of recombinants. Customary units cannot be used because we cannot directly visualize genes with the light microscope. However, due to the relationship between distance and crossover frequency, we may use the map unit. ANALYSIS QUESTIONS: Complete these following the lab. Using the data in Table 1, determine the distance between the gene for spore color and the centromere. Calculate the percent of crossovers by dividing the number of crossover asci (2:2:2:2 or 2:4:2) by the total number of asci x 100. To calculate the map distance, divide the percentage of crossover asci by 2. The percentage of crossover asci is divided by 2 because only half of the spores in each ascus are the result of a crossover event (Figure 3). Record your results in Table 2. Percent crossovers = crossover asci/ total asci X 100 Show how you would get a 2:4:2 arrangement of the ascospores by crossing over. Refer to the figure that shows the formation of crossover asci on page 6; however, do not just copy this figure because it shows specifically how a 2:2:2:2 asci arrangement occurs.