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.

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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

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UF Floriculture Biotech

• Development
• Phase 1 1995-2000 (then)
• Biotechnology
• Phase 2 2000-2005 (now)
• Functional Genomics
• Phase 3 2005-2010 (next)
• Gene Function

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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

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Phase 1 1995-2000

• Transgenic Plants - hormones
• Gibberellins
• Dwarf plants
• Cytokinins
• Stay Green leaves
• Ethylene
• Long lasting flowers

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Internal Growth Regulators
AtGA2ox7 Makes dwarf tobacco
Schomburg et al. 2003
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Internal Growth Regulators
AtGA2ox7 - Makes dwarf petunia
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Stay Green Leaves
sag-IPT Makes leaves stay green longer in
tobacco
Gan Amasino 1995
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sag-IPT
Makes leaves stay greener longer in petunia
Clark et al. in press
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sag-IPT Drought Tolerance
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Long Lasting Flowers
Etr1-1
ETR1-1 - Makes plants ethylene insensitive
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Long Lasting Flowers
Control
Etr1-1
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Problems with 35Setr1-1

• etr1-1 petunias commercially limited

rooting
pathogens etr1-1
MD
etr1-1
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Conclusions Phase 1

• We have a good experimental system
• We have good people in the lab
• We have good collaborators
• We need different genes
• We need better promoters

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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

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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

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Petunia EST Project (EST Expressed Sequence
Tags)
mRNA extraction
Tissue collection
Bioinformatics and Microarrays
library synthesis
EST sequencing
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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

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Petunia EST Database
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Microarray Construction
4200 unique petunia genes
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Microarray analysis Ethylene Regulated Genes
Green AgtB Yellow AB Red AltB
Control (A)
Ethylene (B)
AB
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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

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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

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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

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Measuring Floral Scent
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Flower Scent Composition - Petunia
Benzoic acid carboxyl methyltransferase (BAMT) -
makes methyl benzoate in petunia
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Stop and Smell the Roses!
US consumers bought almost 130 million roses on
Valentines Day 2003 US consumers spent over
7.5B on cut flowers in 2002
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Opportunity Knocks!

• Commercial cut rose varieties dont have much
  fragrance

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Flower Scent Composition - Rose
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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

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Who supports us?

• Financially
• USDA Floriculture Initiative
• American Floral Endowment
• Fred Gloeckner Foundation
• Florida FIRST
• The Scotts Company

• Intellectually
• Harry Klee UF
• Bill Farmerie UF
• Don McCarty UF
• Michelle Jones OSU
• Jim Giovannoni USDA/Cornell
• EU PetNet Group

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UF Floriculture Biotechnology2002-2003
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Questions?