Agriculture Reference
In-Depth Information
primary structure (the sequence); a gene
occupies a particular location in the genome;
a gene encodes an RNA with a particular
expression pattern; genes are coding for a
protein or an mRNA with a dei ned function.
h is means that information on a gene can
be collected at various points during
development using dif erent experimental
approaches.
h e discovery of the gene corresponding
to the desired trait is the i rst step. In the
context of developing transgenic crops, the
choice of traits (genes) to be introduced will
either be problem driven or opportunity
driven. h e i rst category is
mainly input
traits such as herbicide resistance, resistance
to abiotic stresses and pest resistance. h e
second group depends on output traits,
resulting in crops of which the harvested
material has either new or improved
characteristics. In the early days of plant
genetic engineering, the capacity to identify
and isolate the gene of interest was a
signii cant restriction in applying transgene
technology for crop improvement. h e
source of genes that were available for
genetic engineering was rather limited.
Often, these genes were discovered by
accident. Many of the genes used were
isolated from bacteria, such as antibiotic-
resistance genes, herbicide-resistance genes
or genes coding for insecticidal proteins.
Later, due to the huge and exponential
ef orts in plant molecular biology research, a
broad range of genes, also of plant origin,
became available. Important to the discovery
and characterization of these genes was the
introduction of
A. thaliana
as a model plant.
h e assembly of gene libraries and the
characterization of the function of isolated
genes using tools such as insertion
mutagenesis resulted in a broad range of
mutants of which it was known which trait
was af ected. Subsequently, mapping and
sequencing of the mutant genomes allowed
the identii cation of the gene that was
encoding the trait of interest. Nowadays, the
possibilities for discovering genes coding for
particular traits is substantially increased by
the introduction of bioinformatics and
DNA sequencing, especially next-generation
sequenc ing.
h e main source of empirical information
about gene function and structure has been,
and still is, the sequencing of mRNA
transcripts and corresponding cDNAs.
Especially, the use of high-throughput
methods applied on model plants, both
monocots and dicots, has been very
important. In this case, the DNA sequence is
the starting point, and methods such as
protein purii cation, complementation of
mutant phenotypes and reversed genetics
are examples of experimental approaches in
identifying a gene's function. However, for
50% of the genes of most organisms, the
functional or physiological properties of a
gene product are still unknown. Recent
improvements in sequencing methods have
made it possible to generate huge data sets
on the DNA sequences of whole genomes. In
combination with the development of
bioinformatic tools, new opportunities for
identifying genes through exon discovery in
genomic sequence data have been created.
Two approaches can be used to identify
candidate genes: making use of previously
described gene sequences in other species or
starting from the intrinsic characteristic of
genes (e.g. nucleotide composition and
sequence motifs).
h us, as a result of fundamental research
in the domain of gene function analysis over
the last decades, enormous progress has
been made in understanding how thousands
of gene products interact with each other,
resulting in a functional organism of
which the developmental processes are co-
ordinated. It has also provided insight on
how an organism has the capacity to react
towards environmental challenges.
h e research output with the model
species, Arabidopsis, allows the use of genes
as such for genetic engineering, but it is also
a bridge to isolate the homologous genes
from the crop species. Molecular methods
making use of cDNA libraries, gene libraries
and the introduction of PCR technology
have speeded up this process of isolating
genes from crop plants on the basis of the
knowledge obtained from the model species,
Arabidopsis.
By the introduction of new analytical
methods, such as metabolomic proi ling and
