tetrad analysis - fungal genetics

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Tetrad Analysis - Fungal Genetics

Ascomycetes - meiotic spores in ascusEx. Sordaria, Neurospora, Saccharomyces

perithecium

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Tetrad Analysis - Fungal Genetics

Haploid - mycelia or single cells

Tetrad

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Life Cycle of Saccharomyces cerevisiae

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Unordered Tetrads

Saccharomyces cerevisiae

From Al Kapp’s comic ‘Li’l Abner’ 1943

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Unordered Tetrads

Saccharomyces cerevisiae

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Ordered Tetrads

Sordaria, Neurospora

Each chromatid- one spore

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Ordered Tetrads

Sordaria, Neurospora

Each cell from meiosis isrepresented by two identical spores

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Tetrad Analysis

Evaluate by direct examination or tetrad dissection

Sordaria fimicola

tan, black gray, white

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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.

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Determining Number of Genes Involved

Parents crossed: tan x gray # genes for color?

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Ascus Types Expected - Independent Assortment

If Unlinked,

PD = NPD > TT

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Ascus Types Expected - Linked Genes

PD

TT

PD

More frequent, fewer COs

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Ascus Types Expected - Linked Genes

TT

TT

NPD

If linked,

PD >> NPD Less frequent, more COs

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Determining MU Between Linked Genes

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Distance Between Genes - Sample Problem

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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

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Distance Between Gene and Centromere

Only possible to determine with Ordered Tetrads

Distance Gene to Cen = 1/2 (# MII asci) x 100 total

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Distance Gene and CEN - Sample Problem

Where did thecrossovers occur?

Distance:

Aa to Cen?

Bb to Cen?

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Fungal Genetics - Sample Problem

ClassifyPD, TT, NPD

Linked?PD>>NPD?

Distance:

Kk - Ll

Kk and Cen?

Ll and Cen?

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Determining Gene Order

MII means CO has occurred between gene and centromere

Relative frequency: Lowest DCO; Intermediate SCO

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Determining Gene Order

Possible Orders COs required forMII both genes

2 CO

1 CO

1 CO

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Determining Gene Order

When MII for both is lowest frequency, genes are on opposite sides.

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Determining Gene Order

When MII for both is intermediate, genes are on the same side.

Which gene is farthest away?

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Determining Gene Order

MII for both genesIntermediate frequency

MI for inner, MII for outerIntermediate frequency

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Determining Gene Order - Sample Problem

Number of CO?

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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.

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Ordered Tetrad Analysis - Tracking Crossovers

Where did crossoversoccur?

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Mechanism - Homologous Recombination

Point to point pairing, breakage and rejoining

How does this occur?

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Mechanism - Homologous Recombination

Model must explain:

1) Conversion Asci

2) CO intermediates observed

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Holliday Model - Homologous Recombination

First step,nicks in backbones of two ds DNAs

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Holliday Model - Homologous Recombination

Exchange strands

Strand migration

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Holliday Model - Resolution of Chiasma

Nick Outer Strands Cross Over Obvious

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Holliday Model - Resolution of Chiasma

Nick Inner Strands No Cross Over Observed

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Holliday Model - Resolution of Heteroduplexes

Mispairing within heteroduplexes (hybrid regions)

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Holliday Model - Resolution of Heteroduplexes

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Holliday Model - Overview in Text

Box 15.1Russell, 2008Available On Line