Rules: Cell phones off Computers only for class related work No food or drink in lab room Text Book: Web page: Biol 163 Laboratories in Genetics

Goals for course: Reinforce basic genetic principles Introduce model organisms commonly used by geneticists Learn how genetics is used to understand Disease Biochemical pathways Development

Lab reports: Abstract Introduction Results Discussion

Grading: Lab Reports: 50% of grade 5% of that is participation 1 day late, 50% off more than that will only be graded under special circumstances. Research Paper: 10% of grade Outline due Oct. 31. Paper due Dec. 5 2 quizzes: 10% each of final grade. Oct. 3 and Nov. 7. Final exam: 20% of final grade comprehensive Dec. 10.

Genes Alleles and Epistasis

Genetics starts with observation Use genetics to understand the cause of the variability. What proteins or RNAs are responsible for the variability you can see? Observe variability

Easy example, flower color

First make sure the types are heritable and true breeding (homozygous for flower color alleles) X Red by red All uniform

What are the relationships between color types? Red is dominant to Stripes X

Red is dominant to White X

Striped is dominant to White (in some cases) X

We have three types of colored flowers. What does this tell us about flower color? Red is dominant: The plants with red flowers have productive alleles at all genes necessary and sufficient to make red pigment. White is recessive: White flowered plants are defective in some protein or RNA message needed for producing red pigment. Stipes is a plant with 2 alleles of a gene necessary for red pigment. The DNA encoding those genetic variants is sufficiently different from the red allele that only some cells in the petals can make red pigment.

Striped is not always dominant to white. White 2Striped Red X RRa v a v rrAA RrAa v More than one gene is needed for red pigment. Progeny has a functional allele for each gene. Parents each were homozygous for mutant alleles but for different genes required for red pigment.

Complementation test distinguishes if one gene or two affects color One gene aa Two genes RrAa X White 1 White X aa RRaa rrAA White Y Note: the Striped allele is a variant of gene A It is allelic to the mutation in White 1 (RRa v a v)

Complementation tests can be made between recessive alleles. A dominant allele cannot be used. Why?

Yeast complementation test for this week: Brewers Yeast Saccharomyces cerevisiae: 16 chromosomes 12,052 kb DNA 6183 ORFs About 5800 expected to encode proteins

Yeast is a very useful model for genetics because of its life cycle Haploid life cycle

Yeast is a very useful model for genetics because of its life cycle Mating cycle Diploid

We can isolate mutants as haploids We can test the mutations for allelism by a complementation test Two haploids are mated. The resulting Diploid has both mutations. Either the mutations are allelic and do not complement, or they are mutations in two different genes and they do complement.

a1 a2 Select mutants that are defective in Adenine synthesis- cannot grow without adenine in medium. Turn red on media with adenine because an adenine precursor accumulates. a1

a2 a1 X X a2 Which mating results in complementation?

Epistasis Two genes for flower color Two steps in a pathway to make pigment Where are the two genes in the pathway?

Two genes that when mutant lead to white Two genes Rr and Aa RrAa White 1 rrAA White 2 RRaa X

White 1 RRaa Striped is dominant to White 1 RRa v a v RRaa v X

F1 RRaa v F2 3RRa v - 1RRaa X and Striped is allelic to White1 Striped RRa v a v

Punnet square Ra v Ra Ra v Ra RRa v a v RRa v a RRaa Female gametes Male gametes 3RRa v _to 1RRaa

Chi-square test to determine if results support predictions: Hypothesis: striped and white segregate 3:1 From 40 plants, 32 striped and 8 white Chi-square: (O 1 -E 1 ) 2 /E 1 + (O 2 -E 2 ) 2 /E 2 : df = n-1 (30-32) 2 /30 + (10-8) 2 /10 = 4/30+4/10 =.53

Epistasis determines the order of gene products in a pathway. If a plant has two not fully functional (recessive) allele at each of two genes necessary for pigment production, what is the phenotype of the plant? That phenotype tells which gene is first in a pathway. We need to compare recessive alleles that cause distinguishable phenotypes. Eg Striped is a v a v and white 2 is rr What is the petal color of rra v a v ?

Striped and White2 complement White 2 rrAA Striped RRa v a v Red RrAa v X

Alleles at two genes segregate 9:3:3:1 9 R_A_ 3 R_a v 3 rrAA1rra v a v F1 RrAa v X ?

Punnet Square: two genes with randomly segregating alleles Male gametes Female gametes RA Ra v rA ra v rARa v RA RRAA RRAa v RrAARrAa v RRa v a v RrAa v Rra v a v RrAARrAa v rrAA rra v a v RrAa v Rra v a v rrAa v 9R_A_3R_a v a v 3rrA_ 1rra v a v RrAa v X RrAa v

Epistasis tells us the order of genes in a pathway 9 red R_A_ 3 Striped R_a v 4 white R is epistatic to A, R is above A in a pathway F1 RrAa v X

Anthocyanin Pathway: R regulatory gene transcription factor needed to express A protein. The A protein is required for synthesis of pigment It could also be that R produced a precursor chemical needed for A to process into pigment. If there is no R product, no A product can be made. You need the product of R in order to make the product of A. If you have no R, you get R phenotype no matter what is at A. In recessive epistasis this generally means R is upstream of A.

Lab experiment: Corn kernels R and P RR or Rr is full color rr is no color PP or Pp are full color pp is weak color Question is pprr weak color? P epistatic to R or Is pprr no color? R epistatic to P

End

White 2 rrAA Red is dominant to white 2 RRAA RrAA x

Punnet Square: two genes with segregating alleles Male gametes Female gametes RA Ra rA ra rARaRA RRAA RRAa RrAA RrAa RRaaRrAa Rraa RrAA RrAarrAA rraaRrAA Rraa rrAa 9R_A_3R_Aa3rrA_1rraa

9 R_A_ 3 R_aa 3 rrAA3rraa RrAa x