Lecture 2 In this lecture we are going to consider experiments on yeast a very useful organism for genetic study Yeast is more properly known as Saccharomyces cerevisiae which is the single celled microbe used to make bread and beer Yeast can exist as haploids of either a Haploid cells of different mating type mating type MAT or mating type a MATa when mixed together will mate to make a diploid cell Haploids and diploids are isomorphic meaning that a given mutation will cause essentially the same change in haploid and diploid cells This allows us to look at the effect of having two different alleles in the same diploid cell All yeast needs to grow are salts minerals and glucose minimal medium From these compounds yeast cells can synthesize all of the molecules such as amino acids and nucleotides that are needed to construct a cell The synthesis of complicated molecules requires many enzymatic steps When combined these enzymatic reactions constitute a biochemical pathway Consider the pathway for the synthesis of the amino acid histidine A B C D histidine Protein 2 3 4 Enzyme 1 Each intermediate compound in the pathway is converted to the next by an enzyme For example if there is a mutation in the gene for enzyme 3 then intermediate C can not be converted to D and the cell can not make histidine Such a mutant will only grow if histidine is provided in the growth medium This type of mutation is known as an auxotrophic mutation and is very useful for genetic analysis growth on minimal growth on minimal histidine His wild type His Phenotype All traits of an organism with an emphasis on trait under investigation Homozygote diploid with two like alleles of same gene Heterozygote diploid with two different alleles of same gene Recessive Allele trait not expressed in heterozygote genotype phenotype Mate to diploid genotype diploid phenotype MATa a His3 His MAT His3 His3 His3 His MATa a His3 His MAT His3 His3 His3 His Based on the His phenotype of the His3 His3 heterozygote we would say that His3 is recessive to wild type Let s consider a different kind of mutation giving resistance to copper that occurs in a gene known as CUP1 genotype phenotype MATa a Cup1r copper resistant Mate to diploid genotype diploid phenotype MAT Cup1 Cup1r Cup1 copper resistant Dominant Allele trait is expressed in heterozygote Cup1r is dominant to wild type Cup1 The terms dominant and recessive are simply shorthand expressions for the results of particular experiments If someone says a particular allele is dominant that means that at some point they constructed a heterozygous diploid and found that the trait was expressed in that diploid Note Sometimes an allele will have more than one phenotype and may be recessive for one and dominant for another In such cases the phenotype must be specified when one is making statements about whether the allele is dominant or recessive Consider for example the allele for sickle cell hemoglobin in humans designated Hbs Heterozygous individuals Hbs Hba are more resistant to malaria thus Hbs is dominant for the trait of malaria resistance On the other hand Hbs Hba heterozygotes do not the debilitating sickle cell disease but Hbs Hbs homozygous individuals do Therefore Hbs is recessive for the trait of sickle cell disease Once we find out whether an allele is dominant or recessive we can already infer important information about the nature of the allele The following conclusions will usually be true Recessive alleles usually cause the loss of something that is made in wild type Dominant alleles usually cause increased activity or new activity It turns out that the Cupr allele actually carries more copies of the gene for a copper binding protein and therefore increases the activity of the gene Last lecture we defined the gene structurally as the DNA needed to encode a protein We can now define a gene in a new way based on its function Using the phenotypic difference between wild type and a recessive allele we can use a Complementation test to determine whether two different recessive alleles are in the same gene Say you isolate a new recessive histidine requiring mutation that we will call HisX In principle this mutation could be in His3 or it could be in any of the other genes in the histidine biosynthetic pathway In order to distinguish these possibilities we need a test to determine whether HisX the same as His3 To carry out a complementation test one simply constructs a diploid carrying both the His3 and HisX alleles a His3 strain An easy way to do this would be to mate a MAT HisX strain to a MATa possibility tion genotype of diploid phenotype of diploid complementa HisX His3 His3 His3 His No HisX His3 His3 His3 HisX HisX His Yes Having performed this test if the two mutations don t complement we conclude that they are in the same gene Conversely if they do complement we conclude that they are in different genes This test only works for recessive mutations Think about what the outcome would be if HisX were dominant The complementation test can be thought of in the following way If I have an allele with an observable phenotype whose function can be provided by a wild type genotype i e the allele is recessive I can ask whether the function that was lost because of the recessive allele can be provided by another mutant genotype If not the two alleles must be defective in the same gene The beauty of this test is that the trait can serve as a read out of gene function even without knowledge of what the gene is doing at a molecular level Definitions from the Language of Genetics Gene The fundamental unit of heredity which can be defined in three ways i A gene can be defined in molecular terms as a segment of DNA carrying the information necessary to express a complete protein or RNA molecule including the promoter and coding sequence ii A gene can be defined by function with a group of recessive mutations that do not complement each other iii A gene can be defined by position with a single locus segregation pattern in a cross between lines with different alleles Examples are a 1 3 phenotypic ratio in the F2 generation in a cross between diploid organisms or a 2 2 segregation pattern in yeast tetrad analysis Alleles Distinguishable versions of the same gene Locus The site on a chromosome where a gene is located Usually defined by recombinational mapping relative to neighboring loci Genotype The allelic constitution of an individual usually with emphasis on the gene or genes under
View Full Document
Unlocking...