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The Structure and Function of the Expressed Portion of the Wheat Genomes

The Structure and Function of the Expressed Portion of the Wheat Genomes

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The Structure and Function of the Expressed Portion of the Wheat Genomes. The Structure and Function of the Expressed Portion of the Wheat Genomes. The long term goal of this project is to decipher the chromosomal location and biological function of all genes in the wheat genomes. - PowerPoint PPT Presentation

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Page 1: The Structure and Function of the Expressed Portion of the Wheat Genomes

The Structure and Function of the

Expressed Portion of the Wheat Genomes

Page 2: The Structure and Function of the Expressed Portion of the Wheat Genomes

The Structure and Function of the

Expressed Portion of the Wheat Genomes

The long term goal of this project is to decipher the chromosomal location and biological

function of all genes in the wheat genomes.

Page 3: The Structure and Function of the Expressed Portion of the Wheat Genomes

The Structure and Function of the Expressed Portion of the Wheat Genomes

Value and uses of wheat

Worldwide, wheat (bread and durum) is

the most widely grown crop.

With its high protein content, wheat is the single most important source of plant protein

in the human diet.

In the USA, wheat is the fourth most economically

significant crop and the leading export

crop.

Wheat products: seed

flourbeverageglutenstarch

Page 4: The Structure and Function of the Expressed Portion of the Wheat Genomes
Page 5: The Structure and Function of the Expressed Portion of the Wheat Genomes

Cal Qualset, U Calif DANR, DavisJames Anderson, U MinnesotaOlin Anderson, U Calif DANR, AlbanyTimothy Close, U Calif RiversideJorge Dubcovsky, U Calif DavisJan Dvorak, U Calif DavisBikram Gill, Kansas State UKulvinder Gill, U NebraskaJ Perry Gustafson, U MissouriShahryar Kianian, N Dakota State UNora Lapitan, Colorado State UHenry Nguyen, Texas Tech UMark Sorrells, Cornell UM Kay Walker-Simmons, Washington State U

Page 6: The Structure and Function of the Expressed Portion of the Wheat Genomes

1. Produce cDNA libraries from as many tissue and condition combinations as possible.

Objectives and Experimental Approach

Produce multiple cDNA libraries with a target of 30 total after quality testing, normalizing and subtraction to reduce redundancy.

Page 7: The Structure and Function of the Expressed Portion of the Wheat Genomes

1. Produce cDNA libraries from as many tissue and condition combinations as possible.

Objectives and Experimental Approach

2. Determine the base-pair sequence of these cDNAs, yielding ESTs.

Carry out 5’end sequencing of cDNA libraries, with 3’ sequencing of putative singletons.

Produce multiple cDNA libraries with a target of 30 total after quality testing, normalizing and subtraction to reduce redundancy.

Page 8: The Structure and Function of the Expressed Portion of the Wheat Genomes

1. Produce cDNA libraries from as many tissue and condition combinations as possible.

Objectives and Experimental Approach

2. Determine the base-pair sequence of these cDNAs, yielding ESTs.

3. Map into wheat deletion stocks a set of 10,000 unique ESTs.

Map ESTs into bins defined by wheat deletion stocks; target is 10,000 mapped ESTs.

Carry out 5’end sequencing of cDNA libraries, with 3’ sequencing of putative singletons.

Produce multiple cDNA libraries with a target of 30 total after quality testing, normalizing and subtraction to reduce redundancy.

Page 9: The Structure and Function of the Expressed Portion of the Wheat Genomes

1. Produce cDNA libraries from as many tissue and condition combinations as possible.

Objectives and Experimental Approach

2. Determine the base-pair sequence of these cDNAs, yielding ESTs.

3. Map into wheat deletion stocks a set of 10,000 unique ESTs.

4. Determine the expression of the mapped ESTs relative to reproductive biology of wheat.

Using arrays, analyze the expression of the mapped ESTs, focusing on five aspects of wheat reproduction.

Map ESTs into bins defined by wheat deletion stocks; target is 10,000 mapped ESTs.

Carry out 5’end sequencing of cDNA libraries, with 3’ sequencing of putative singletons.

Produce multiple cDNA libraries with a target of 30 total after quality testing, normalizing and subtraction to reduce redundancy.

Page 10: The Structure and Function of the Expressed Portion of the Wheat Genomes

1. Produce cDNA libraries from as many tissue and condition combinations as possible.

Objectives and Experimental Approach

2. Determine the base-pair sequence of these cDNAs, yielding ESTs.

3. Map into wheat deletion stocks a set of 10,000 unique ESTs.

4. Determine the expression of the mapped ESTs relative to reproductive biology of wheat.

5. Process, analyze, and display data accumulated in this project (bioinformatics).

Develop and enhance means to analyze, interpret, and visualize project data (data processing, database modifications, and web page maintenance).

Using arrays, analyze the expression of the mapped ESTs, focusing on five aspects of wheat reproduction.

Map ESTs into bins defined by wheat deletion stocks; target is 10,000 mapped ESTs.

Carry out 5’end sequencing of cDNA libraries, with 3’ sequencing of putative singletons.

Produce multiple cDNA libraries with a target of 30 total after quality testing, normalizing and subtraction to reduce redundancy.

Page 11: The Structure and Function of the Expressed Portion of the Wheat Genomes

1. Produce cDNA libraries from as many tissue and condition combinations as possible.

Objectives and Experimental Approach

2. Determine the base-pair sequence of these cDNAs, yielding ESTs.

3. Map into wheat deletion stocks a set of 10,000 unique ESTs.

4. Determine the expression of the mapped ESTs relative to reproductive biology of wheat.

5. Process, analyze, and display data accumulated in this project (bioinformatics).

6. Analyze gene density and distribution of mapped ESTs and thus genes in the wheat genomes (genome structure and evolution).

Analyze densities and distribution of related genes determined from deletion map locations combined with functionality.

Develop and enhance means to analyze, interpret, and visualize project data (data processing, database modifications, and web page maintenance).

Using arrays, analyze the expression of the mapped ESTs, focusing on five aspects of wheat reproduction.

Map ESTs into bins defined by wheat deletion stocks; target is 10,000 mapped ESTs.

Carry out 5’end sequencing of cDNA libraries, with 3’ sequencing of putative singletons.

Produce multiple cDNA libraries with a target of 30 total after quality testing, normalizing and subtraction to reduce redundancy.

Page 12: The Structure and Function of the Expressed Portion of the Wheat Genomes

EST Arrays

SAGE

SequenceMatching

DeletionMapping

ComparativeMapping

Objs. 1 & 2.EST Production

cDNA librariesScreening/normalizationsSequencingData analysisDNA storage/distribution

Obj. 6.Genome

Structure &Evolution

Obj. 3.Mapping

Obj. 4.FunctionalGenomics

The Structure and Function of the Expressed Portion of the Wheat Genomes

Page 13: The Structure and Function of the Expressed Portion of the Wheat Genomes

Project Coordinator: Calvin QualsetProject Manager: Patrick McGuire

Objectives 1 and 2.EST Production

Coordinator:Olin Anderson

Objective 5.Bioinformatics

Coordinator:Olin Anderson

Objective 3.Mapping

Coordinator:Bikram Gill

Objective 4.Functional Genomics

Coordinator:Mark Sorrells

Objective 6.Genome Structure & Evolution

Coordinator:Jan Dvorák

EST Arrays

SAGE

SequenceMatching

DeletionMapping

ComparativeMapping

Objs. 1 & 2.EST Production

cDNA librariesScreening/normalizationsSequencingData analysisDNA storage/distribution

Obj. 6.Genome

Structure &Evolution

Obj. 3.Mapping

Obj. 4.FunctionalGenomics

The Structure and Function of the Expressed Portion of the Wheat Genomes

Page 14: The Structure and Function of the Expressed Portion of the Wheat Genomes

Use of chromosome deletion stocksExample: chromosome 6B

6B

In this physical model of the chromosome, the dark areas are the heterochromatic bands which serve as landmark patterns identifying this chromosome uniquely. The arrows indicate breakpoints of the deletion stocks available.

#1 #2 #3

Page 15: The Structure and Function of the Expressed Portion of the Wheat Genomes

Use of chromosome deletion stocksExample: chromosome 6B

6B

In this physical model of the chromosome, the dark areas are the heterochromatic bands which serve as landmark patterns identifying this chromosome uniquely. The arrows indicate breakpoints of the deletion stocks available.

#1 #2 #3

If probe X produces a signal in this stock, ...

Page 16: The Structure and Function of the Expressed Portion of the Wheat Genomes

Use of chromosome deletion stocksExample: chromosome 6B

6B

In this physical model of the chromosome, the dark areas are the heterochromatic bands which serve as landmark patterns identifying this chromosome uniquely. The arrows indicate breakpoints of the deletion stocks available.

#1 #2 #3

… and in this stock, ...

If probe X produces a signal in this stock, ...

Page 17: The Structure and Function of the Expressed Portion of the Wheat Genomes

Use of chromosome deletion stocksExample: chromosome 6B

6B

In this physical model of the chromosome, the dark areas are the heterochromatic bands which serve as landmark patterns identifying this chromosome uniquely. The arrows indicate breakpoints of the deletion stocks available.

#1 #2 #3

… and in this stock, ...

… but not in this stock, then ...

If probe X produces a signal in this stock, ...

Page 18: The Structure and Function of the Expressed Portion of the Wheat Genomes

Use of chromosome deletion stocksExample: chromosome 6B

6B

In this physical model of the chromosome, the dark areas are the heterochromatic bands which serve as landmark patterns identifying this chromosome uniquely. The arrows indicate breakpoints of the deletion stocks available.

#1 #2 #3

… and in this stock, ...

… but not in this stock, then ...

… one concludes that the DNA sequence in this chromosome corresponding to the EST represented by probe X is physically located in this region or ‘bin’.

If probe X produces a signal in this stock, ...

Page 19: The Structure and Function of the Expressed Portion of the Wheat Genomes

Use of chromosome deletion stocksExample: chromosome 6B

6B

In this physical model of the chromosome, the dark areas are the heterochromatic bands which serve as landmark patterns identifying this chromosome uniquely. The arrows indicate breakpoints of the deletion stocks available.

Cumulatively, these deletion stocks define ‘bins’ of various sizes in the physical chromosomes. The full 10,000 ESTs

developed in this project will be mapped into these bins.

#1 #2 #3

… and in this stock, ...

… but not in this stock, then ...

… one concludes that the DNA sequence in this chromosome corresponding to the EST represented by probe X is physically located in this region or ‘bin’.

If probe X produces a signal in this stock, ...

Page 20: The Structure and Function of the Expressed Portion of the Wheat Genomes

Functional Genomics: five aspects of wheat reproduction

The Structure and Function of the Expressed Portion of the Wheat Genomes

Flowering Signals

Meiosis

Pollen Development

Seed DevelopmentDormancy & Germination

Page 21: The Structure and Function of the Expressed Portion of the Wheat Genomes

Deliverables

30 cDNA libraries, normalized and enhanced for abundance of different sequences

Sequence database for 10,000 singleton ESTs for bread wheat

Physical bin map of the singleton ESTs in the wheat genomes

Arrays of the mapped singleton ESTs for gene discovery efforts

Expression profiles for wheat genes during five stages of reproduction: flowering initiation; meiosis; pollen development and nucleo-cytoplasmic interactions; seed development; and seed dormancy/germination

Informatics sites and software for sequence analyses, comparisons, and display and for differential display analyses of probed arrays

A minimum of 13 young professionals with training and experience with tools and technology of functional genomics

A nucleus of 13 public labs equipped, experienced, and networked for gene discovery and deployment and training with wheat or other species

Page 22: The Structure and Function of the Expressed Portion of the Wheat Genomes

The Structure and Function of the Expressed Portion of the Wheat Genomes

Biology of bread wheat

Triticum aestivum L., tribe Triticeae, family Poaceae

Allohexaploid: six component genomes, AABBDD, 2n=6x=42

Bread wheat originated from two hybridization events, the first between two diploid species producing an AABB genome tetraploid, the second between the tetraploid species and another diploid species

Closest relatives to bread wheat in the tribe:the crops durum wheat (tetraploid, AABB), einkorn wheat (diploid, AA), barley (HH), rye (RR), and triticale (AABBRR), numerous weedy annual species of Aegilops and Triticum, and numerous forage species

Diploid species are the sources of the wheat genomes: A from Triticum urartu B from a primitive form of Aegilops speltoides or an extinct close relative D from Aegilops tauschii

The molecular genetic map has some 2000 markers.

Each chromosome pair has a unique pattern of heterochromatic bands allowing unequivocal cytological identification

Wheat tolerates aneuploidy and a huge library of aneuploid stocks exists in many different genotypes. Monosomics Deletions Ditelosomics Translocations Nullisomics-tetrasomics Alien introgression stocks

Aneuploids have been critical in gene discovery, gene mapping, chromosome manipulation, and gene transfer in classical cytogenetics and breeding. Coupled with molecular technology, the aneuploids offer tools for greater progress in all these areas plus gene cloning, transformation, and physical mapping.

Aneuploids have facilitated the understanding of syntenic relationships among thee three genomes: seven homoeologous groups have been defined, each with a chromosome from each genome.