1

Tetrad Analysis - Fungal Genetics

Ascomycetes - meiotic spores in ascusEx. 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 isrepresented 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

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 betweenGene 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 thecrossovers occur?

Distance:

Aa to Cen?

Bb to Cen?

19

Fungal Genetics - Sample Problem

ClassifyPD, 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 Orders COs required forMII both genes

2 CO

1 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 genesIntermediate frequency

MI for inner, MII for outerIntermediate 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 crossoversoccur?

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 15.1Russell, 2008Available On Line