Hunting For Mutants In Saccharomyces Cerevisiae
David Kennedy
Bio 260
Section 301
6/5/13
Hunting for Mutants in Saccharomyces cerevisiae
Purpose: This experiment was conducted in order to understand patterns of inheritance through mutations that occur to the model organism Saccharomyces cerevisiae (budding yeast). The experiment found auxotrophic and temperature sensitive mutants. Auxotrophic mutants are mutants that have defect(s) in one or more genes involved in biosynthetic pathways. Temperature sensitive mutants in this experiment, are mutants that do not survive at higher (restrictive) temperatures. This experiment compared mutant colonies with a control group.
Experimental Design:
Week 1:
One 1ml aliquot of mutagenized yeast culture in buffered saline solution was utilized to begin the experiment. The yeast culture in saline solution was shaken, in order to suspend the yeast throughout the solution. 10 micro liters of the solution was taken and added to 90 micro liters of water, in order to be able to count the number of cells through a hemocytometer. A Pasteur pipette was used to pipette a sample of the diluted cell culture into the hemocytometer. The cells on the small hemocytometer grid was counted, in order to estimate the number of cells that were exposed to the mutagen.
The cells were further diluted in order to obtain individual colonies to be counted later, or there would be an uncountable “carpet” of yeast on agar plates. 0.1 mL of cell culture was pipetted into a 9.9 mL water blank. This was then mixed thoroughly (to suspend all yeast cells), and a fresh pipette tip was used to transfer 0.1 mL of this solution into a new 9.9 mL water blank. This was mixed again and 1.0 mL of this solution was pipetted into 9.0 mL water blank (to achieve a final dilution of 1:100,000 for the agar plates).
3 YPD media agar plates were obtained and labeled with group number and the date. Each plate was also labeled 0.1 mL, 0.2 mL, 0.4 mL, these corresponded to the volume of mutated cell culture
Bio 260
Section 301
6/5/13
Hunting for Mutants in Saccharomyces cerevisiae
Purpose: This experiment was conducted in order to understand patterns of inheritance through mutations that occur to the model organism Saccharomyces cerevisiae (budding yeast). The experiment found auxotrophic and temperature sensitive mutants. Auxotrophic mutants are mutants that have defect(s) in one or more genes involved in biosynthetic pathways. Temperature sensitive mutants in this experiment, are mutants that do not survive at higher (restrictive) temperatures. This experiment compared mutant colonies with a control group.
Experimental Design:
Week 1:
One 1ml aliquot of mutagenized yeast culture in buffered saline solution was utilized to begin the experiment. The yeast culture in saline solution was shaken, in order to suspend the yeast throughout the solution. 10 micro liters of the solution was taken and added to 90 micro liters of water, in order to be able to count the number of cells through a hemocytometer. A Pasteur pipette was used to pipette a sample of the diluted cell culture into the hemocytometer. The cells on the small hemocytometer grid was counted, in order to estimate the number of cells that were exposed to the mutagen.
The cells were further diluted in order to obtain individual colonies to be counted later, or there would be an uncountable “carpet” of yeast on agar plates. 0.1 mL of cell culture was pipetted into a 9.9 mL water blank. This was then mixed thoroughly (to suspend all yeast cells), and a fresh pipette tip was used to transfer 0.1 mL of this solution into a new 9.9 mL water blank. This was mixed again and 1.0 mL of this solution was pipetted into 9.0 mL water blank (to achieve a final dilution of 1:100,000 for the agar plates).
3 YPD media agar plates were obtained and labeled with group number and the date. Each plate was also labeled 0.1 mL, 0.2 mL, 0.4 mL, these corresponded to the volume of mutated cell culture