Agriculture Reference
In-Depth Information
for the adenosine-5
0
-phosphosulfate reductase, a key enzyme for assimilatory
sulfate reduction pathway. The authors showed that the difference in sulfate
contents between the two parental lines was not due to a different expression of
the gene in the two genotypes, but to a change of alanine into glutamic acid in the
APR2 protein. However, it is possible for there to be no overlap between the QTL
and the candidate genes, as happened in the study on common bean conducted by
Blair et al. (
2010
). A QTL for iron reductase activity in roots was identified under
iron sufficiency (15
g Fe), but it was mapped on a different chromosome from the
one found under iron-limited growth (1
μ
M). Therefore, it was postulated that iron
reductase activity was influenced by more than one locus, with a first iron
reductase-related locus on chromosome b02, which contributed to the trait in
iron-limited plants, and a second iron reductase-related locus present on chromo-
some b11 contributing in iron-sufficient plants. The authors also mapped loci for
the FRO genes (iron-reductase homologues) but since there is no co-localisation
with the QTL they concluded that some other gene may control iron reductase
activity. Another resource for finding candidate genes is analysing gene expression
in the vicinity of the QTL. This can be done using standard assays for a limited
number of candidate genes, or using high-throughput genome-wide techniques,
such as microarrays (Borevitz and Nordborg
2003
). When the functional allelic
variation results in gene expression differences, this may clearly indicate the
candidate gene. Identifying an artificially induced mutant showing phenotypic
effect in the trait of interest provides a unique functional argument to select a
candidate gene. The availability of T-DNA insertion mutants for almost any
Arabidopsis gene and the efficiency of TILLING (Targeting Induced Local Lesions
in Genomes) procedures to identify mutations in numerous candidate genes provide
efficient strategies to analyse knock-out phenotypes of (nearly) all genes in a QTL
region. Nevertheless, most collections of mutants are in the laboratory backgrounds
L
er
(Landsberg
erecta
) and Col (Columbia), which do not necessarily carry func-
tional alleles at the gene of interest and, consequently, will not always show a
distinct phenotype when mutated. Therefore, loss-of-function mutants of particular
lines, such as NILs, carrying alleles different from the common laboratory acces-
sions, can also be induced by mutagenesis with standard chemical or physical
agents. This approach is especially useful when identifying novel alleles that are
dominant over laboratory backgrounds (Koornneef et al.
2004
). Ultimately, the
proof for the identification of a QTL gene should come from complementation
experiments by plant transformation.
μ
QTL Validation
The presence of a QTL is validated when allelic variation of that QTL area has an
effect on the studied trait. The search for polymorphic region-specific markers is
crucial to fine map a QTL. NILs and HIFs are screened with molecular markers that
are contrasted at the target region in parents. Finally, combining the analysis of a
