1 Tetrad Analysis - Fungal Genetics Ascomycetes - meiotic spores in ascus Ex. Sordaria, Neurospora, Saccharomyces perithecium

2 Tetrad Analysis - Fungal Genetics Haploid - mycelia or single cells Tetrad

3 Life Cycle of Saccharomyces cerevisiae

4 Unordered Tetrads Saccharomyces cerevisiae From Al Kapp’s comic ‘Li’l Abner’ 1943

5 Unordered Tetrads Saccharomyces cerevisiae

6 Ordered Tetrads Sordaria, Neurospora Each chromatid - one spore

7 Ordered Tetrads Sordaria, Neurospora Each cell from meiosis is represented by two identical spores

8 Tetrad Analysis Evaluate by direct examination or tetrad dissection Sordaria fimicola tan, black gray, white

9 Classify Ascus ( ordered, unordered ) - Spore Phenotypes Parents crossed: tan x gray t g zygote Parental ditype (PD) also t g t g, g t t g, etc. Nonparental ditype (NPD) also b w w b, b w b w, etc. Tetratype (TT) also b w g t, b t w g, etc.

10 Determining Number of Genes Involved Parents crossed: tan x gray# genes for color?

11 Ascus Types Expected - Independent Assortment If Unlinked, PD = NPD > TT

12 Ascus Types Expected - Linked Genes PD TT PD More frequent, fewer COs

13 Ascus Types Expected - Linked Genes TT NPD If linked, PD >> NPD Less frequent, more COs

14 Determining MU Between Linked Genes

15 Distance Between Genes - Sample Problem

16 Distance Between Gene and Centromere Only possible to determine with Ordered Tetrads MI segregation MI ascus MII segregation MII ascus No CO CO between Gene and Cen

17 Distance Between Gene and Centromere Only possible to determine with Ordered Tetrads Distance Gene to Cen = 1/2 (# MII asci) x 100 total

18 Distance Gene and CEN - Sample Problem Where did the crossovers occur? Distance: Aa to Cen? Bb to Cen?

19 Fungal Genetics - Sample Problem Classify PD, TT, NPD Linked? PD>>NPD? Distance: Kk - Ll Kk and Cen? Ll and Cen?

20 Determining Gene Order MII means CO has occurred between gene and centromere Relative frequency: Lowest DCO; Intermediate SCO

21 Determining Gene Order Possible OrdersCOs required for MII both genes 2 CO 1 CO

22 Determining Gene Order When MII for both is lowest frequency, genes are on opposite sides.

23 Determining Gene Order When MII for both is intermediate, genes are on the same side. Which gene is farthest away?

24 Determining Gene Order MII for both genes Intermediate frequency MI for inner, MII for outer Intermediate frequency

25 Determining Gene Order - Sample Problem Number of CO?

26 Ordered Tetrad Analysis - Sample Problem 1. Classify as PD, NPD, TT. 2. Classify as MI, MII for each gene. 3. Determine linkage, gene order. 4. Calculate distances between genes, each gene and centromere.

27 Ordered Tetrad Analysis - Tracking Crossovers Where did crossovers occur?

28 Mechanism - Homologous Recombination Point to point pairing, breakage and rejoining How does this occur?

29 Mechanism - Homologous Recombination Model must explain: 1) Conversion Asci 2) CO intermediates observed

30 Holliday Model - Homologous Recombination First step, nicks in backbones of two ds DNAs

31 Holliday Model - Homologous Recombination Exchange strands Strand migration

32 Holliday Model - Resolution of Chiasma Nick Outer Strands Cross Over Obvious

33 Holliday Model - Resolution of Chiasma Nick Inner Strands No Cross Over Observed

34 Holliday Model - Resolution of Heteroduplexes Mispairing within heteroduplexes (hybrid regions)

35 Holliday Model - Resolution of Heteroduplexes

36 Holliday Model - Overview in Text Box 15.1 Russell, 2008 Available On Line