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Chapter 11: Functional genomics
Finding genes and their function
Structural genome analysis: find genes
Functional genome analysis: find function
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As we will see in chapter 12, most
transgenic plants that are in the fields
now have been made with bacterial
genes (easier to find and analyse)
For the newer transgenic plants, plant
genes have been introduced.
How can these interesting plant genes be
found?
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Structural genome analysis
Eukaryotic genomes contain much DNA outside the genes
Human
Yeast
Z. mays
E. coli
Genomes 2
T.A. Brown, Bios
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Structural genome analysis
Eukaryotic genomes contain much DNA outside the genes
How to find these genes that are scattered on the genome?
When the genome is sequenced, there are several
techniques to find the genes in the sequence:
• look for coding regions (open reading frames from ATG
to stopcodon), not so easy in eukaryotes because the
open reading frame is interrupted by introns.
• compare DNA with mRNA that is transcribed from the
genes (RNA can be extracted and analysed)
• Structural characteristics (such as GC%)
• Compare with information from other organisms
• Map based cloning: for genes for which a mutant 4
phenotype is known
Map based cloning
• Map based cloning: for genes for which a mutant
phenotype is known
• For this technique, physical and genetic (chapter 9)
maps are needed.
• A physical map is based on a collection of clones
that are overlapping each other and preferentially the
corresponding sequence.
• Such a physical map can be correlated with a
genetic map by comparing the sequence of a genetic
marker with the sequence of the physical map.
• If the mutant phenotype is genetically mapped, than
the gene can be cloned on the basis of the correlated
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physical map = map based cloning
Physical map
Overlapping
clones
DNA
sequence
Genetics, analysis of genes and genomes, Hartl and Jones
(Jones and Bartlett publishers)
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Integration of physical and genetic maps
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Functional genome analysis
• To find the function of the genes
 search for homology with known genes
(chapter 3: BLAST)
 Expression analysis, for example a gene that
is expressed only in the flower must have a
function in the flower
 By mutant analysis
 By making transgenic plants with higher or
lower expression of the gene
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Functional genome analysis
•
By making transgenic plants with much higher
or lower expression of the gene
– Higher expression: by introducing construct with
strong promoter- gene fusion
– Lower expression: with RNAi or antisense technology
Antisense produces the reverse of the normal mRNA
 hybridisation with mRNA prevents translation and
makes mRNA unstable
RNAi uses double stranded RNA and has a similar
effect (complex mechanism)
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Functional genome analysis
•
By making transgenic plants with much higher
or lower expression of the gene, for example a
gene that makes plant grow taller
– Higher expression: transgenic plant will be larger
than normal
– Lower expression: transgenic plant will be smaller
than normal
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ANTISENSE STRATEGY
promoter
DNA
coding
sequence
messenger RNA
protein
coding
promoter sequence
ANTISENSE