Development of cdna microarrays for gene expression research in floriculture crops
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Development of cDNA Microarrays for Gene Expression Research in Floriculture Crops. Dave Clark University of Florida Environmental Horticulture Dept. UF Floriculture Biotech Lab (est. 1995). Our main goal: To bridge the gap between basic molecular biology and applied floriculture

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Development of cDNA Microarrays for Gene Expression Research in Floriculture Crops

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Development of cDNA Microarrays for Gene Expression Research in Floriculture Crops

Dave Clark

University of Florida

Environmental Horticulture Dept.


UF Floriculture Biotech Lab(est. 1995)

  • Our main goal:

  • To bridge the gap between basic molecular biology and applied floriculture

    • Florida produces 20% of US floriculture crops


UF Floriculture Biotech

Development:

  • Phase 1 – 1995-2000 (then)

    • Biotechnology

  • Phase 2 – 2000-2005 (now)

    • Functional Genomics

  • Phase 3 – 2005-2010 (next)

    • Gene Function


Phase 1 – 1995-2000

  • Establish a biotech system

    • Petunia hybrida

  • Proof of concept research

    • Expansion of available technology

  • Build critical personnel mass

    • Recruit good people

    • Make strong collaborations


Phase 1 – 1995-2000

  • Transgenic Plants - hormones

  • Gibberellins

    • Dwarf plants

  • Cytokinins

    • “Stay Green” leaves

  • Ethylene

    • Long lasting flowers


Internal Growth Regulators

AtGA2ox7

Makes dwarf tobacco

Schomburg et al. 2003


Internal Growth Regulators

AtGA2ox7 - Makes dwarf petunia


Stay Green Leaves

sag-IPT

Makes leaves stay green longer in tobacco

Gan & Amasino 1995


sag-IPT

Makes leaves stay greener longer in petunia

Clark et al. in press


sag-IPT – Drought Tolerance


Long Lasting Flowers

Etr1-1

ETR1-1 - Makes plants ethylene insensitive


Long Lasting Flowers

Control

Etr1-1


Problems with 35S::etr1-1

  • etr1-1 petunias – commercially limited

rooting

pathogens – etr1-1

‘MD’

etr1-1


Conclusions – Phase 1

  • We have a good experimental system

  • We have good people in the lab

  • We have good collaborators

  • We needdifferent genes

  • We needbetter promoters


Phase 2 – 2000-2005

  • Functional Genomics

    • New tools for genetics research

  • Exploration for new technologies

    • New and novel genes

  • Controlling gene expression

    • Petal specific promoters


Functional Genomics

  • Tools that help discover the biological function of genes

  • Uncovers how groups of genes work together in a biological process

  • Encompasses many traditional biological and genetic approaches


Petunia EST Project

(EST = Expressed Sequence Tags)

mRNA extraction

Tissue collection

Bioinformatics and Microarrays

library synthesis

EST sequencing


Petunia Flower Genes

  • Petunia gene sequencing:

    Floral development library 2774

    C2H4-treated flower library 3264

    Pollinated flower library 1824

    Other libraries 1046

  • Total genes sequenced 8908


Petunia EST Database


Microarray Construction

4200 unique petunia genes


Microarray analysis – Ethylene Regulated Genes

Green A>B

Yellow A=B

Red A<B

Control

(A)

Ethylene

(B)

A+B


Ethylene Regulated Genes

  • We have isolated approximately 50 genes that are regulated by ethylene

  • Some of these genes are thought to be involved in making floral scent volatiles

    • Benzoic acid carboxyl methyltransferase


Phase 2 – Still going

  • We still bridge the gap between basic molecular biology and applied floriculture

    • We have a petunia EST database

    • Microarrays are working well

    • We have many new and novel genes

    • We have good promoters


Phase 3 – 2005-2010

  • Gene Function

    • We are ahead of schedule:

    • Experimental system (phase 1)

      • Adopted by several groups

    • New genetic tools (phase 2)

      • Available to floriculture researchers

    • Understanding gene function


Measuring Floral Scent


Flower Scent Composition - Petunia

Benzoic acid carboxyl methyltransferase (BAMT) - makes methyl benzoate in petunia


Stop and Smell the Roses!

US consumers bought almost 130 million roses on Valentine’s Day 2003

US consumers spent over $7.5B on cut flowers in 2002


Opportunity Knocks!

  • Commercial cut rose varieties don’t have much fragrance


Flower Scent Composition - Rose


The Future

  • We plan to maintain and grow the genetics database

  • We plan to make transgenic plants expressing new genes

  • We plan to knock out 1 gene a week – functional analysis

  • We plan to continue making new collaborations


Who supports us?

  • Intellectually

    • Harry Klee – UF

    • Bill Farmerie – UF

    • Don McCarty – UF

    • Michelle Jones – OSU

    • Jim Giovannoni – USDA/Cornell

    • EU PetNet Group

  • Financially

    • USDA Floriculture Initiative

    • American Floral Endowment

    • Fred Gloeckner Foundation

    • Florida FIRST

    • The Scotts Company


UF Floriculture Biotechnology2002-2003


Questions?


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