Mapping the genetic loci that control the quantitative variation is a preliminary step to disclose the complex regulation of a polygenic trait. Better knowledge of the genetic determinism of a trait could in turn aid breeders in advancing the crop. Linkage mapping is the traditional method for identifying QTL while association mapping, originally used in humans and animals, has recently been adopted in plants. Association mapping has at least two main advantages over traditional linkage mapping methods (Zhu et al., 2008). First, it mitigates the need to construct population
2.5 QTL mapping for oil and phytosterol content inB. napus 15
from crosses by using natural population which has a much broader germplasm context. Second, it can achieves a higher resolution mapping by exploiting linkage disequilibrium (LD) generated from historical recombination events. In all mapping approaches, however, a trade-off exists between statistical power and resolution. Association mapping is also associated with a higher risk of biased estimation or even false inference due to population structure. As such, an ideal analysis would be to reap the benefits of each method by complementary use of both linkage and association mapping to obtain both high power of detection and resolution.
Over the past few decades, numerous QTL for oil content in oilseed rape have been identified using different mapping methods and different populations. In linkage mapping studies, the segregating populations were developed either from crosses where both parental lines had a high oil content (Zhao et al., 2005), both had a moderate oil content (Ecke et al., 1995; Burns et al., 2003;
Zhao et al., 2005; Qiu et al., 2006; Jiang et al., 2014), or one had a high oil and the other a moderate oil content (Delourme et al., 2006). The first genetic studies which set out to map QTL controlling the seed oil content variation inB. napusdetected three discrete loci (Ecke et al., 1995). Of the three loci, two are closely associated with erucic acid content, indicating a direct effect of the erucic acid genes on oil content. Burns et al. (2003) identified seven QTL using an intervarietal subset of substitution lines. Subsequent study involving a European and a Chinese parental lines ("Sollux"
and "Gaoyou", SG population) detected eight QTL with additive effects and nine pairs of loci with additive×additive epistasis along with high genotype×environment interactions (Zhao et al., 2005). Another similar population generated between a Chinese and a European parental lines ("Tapidor" and "Ningyou7", TN population) identified 7 QTL for oil content in which three were found coincided with QTL for erucic acid (Qiu et al., 2006). Using two populations with different genetic backgrounds ("Darmor-bzhand Yudal", DY population; "Rapid" and "NSL96/25", RNSL population), Delourme et al. (2006) identified 14 and 10 genomic regions associated with seed oil content in which only one QTL was found potentially common to the two populations. The study reported three pairs of epistatic interactions and attributed additive effects as the main factors contributing to variation in oil content.
A larger number of QTL for oil content were reported using association mapping approach. In a first experiment on whole-genome association analysis in oilseed rape, Honsdorf et al. (2010) identified 22 QTL for oil content in a set of 84 canola quality winter. Using a new-type population
and a traditional oilseed rape population, Zou et al. (2010) identified 54 QTL associated with seed oil content, 6 of which were found collocated with QTL detected by Qiu et al. (2006) using interval mapping approach. Another association mapping study which included 17 SNPs derived from 9 candidate genes from the triacylglycerol biosynthetic pathway in a population of 685 diverse elite oilseed rape inbred lines demonstrated that in addition to main effects, both intergenic and intragenic epistasis also contributed a considerable amount of genotypic variation in oil content (Würschum et al., 2013). The identified interactions includes certain key enzymes involved in the main pathway of storage oil formation as well as the WRI1 transcription factor which is known to be involved in the control of storage compound biosynthesis inArabidopsis.
In a recent study, Jiang et al. (2014) updated the number of QTL for seed oil content to 41 in the TN population with increased number of environments and marker density from the previous study reported by Qiu et al. (2006). With an additional TN reconstructed F2 population, Jiang et al. (2014) detected 20 QTL with dominance effects in which a majority of them showed partially dominant effect and only four QTL showed positive complete-dominance or mild over-dominance, suggesting that oil content in oilseed rape has weaker heterosis compared with other traits such as seed yield (Radoev et al., 2008). In an attempt to account for full extent of the variation in seed-oil content, Jiang et al. (2014) also established a reference map by incorporating common markers from different genetic populations (SG, DY and RNSL populations) on the genetic map of TN population. The resulting reference map enabled QTL detected from SG, DY and RNSL populations as well as the significant markers detected by association study of Zou et al. (2010) to be aligned and compared with its own detected QTL. In total, the reference map identified 46 distinct QTL regions that control seed oil content on 16 of the 19 linkage groups ofB. napus. Of the 46 QTL, 18 were identified in multiple populations.
So far, only one QTL mapping study for phytosterol has been reported in oilseed rape (Amar et al., 2008b). By using the population that was previously used by Ecke et al. (1995), Amar et al.
(2008b) reported three QTL for total phytosterol content, two of which were found collocated with erucic acid genes on A08 and C03. Based on the fact that cytoplasmic acetyl-CoA is required as precursor for both synthesis of erucic acid and phytosterols (Figure 2.2) and that the alleles increasing phytosterol content exhibited negative relationship with erucic acid content, the authors further concluded that the two QTL identified for phytosterols were due to pleiotropic effect
2.5 QTL mapping for oil and phytosterol content inB. napus 17
exerted by the two erucic acid genes. As such, it can be anticipated that the utilization of a DH population that does not segregate for erucic acid would lead to increase of detection power for QTL with smaller effects.
Genetic variation and inheritance of phytosterol and oil content in a
doubled haploid population derived from the winter oilseed rape
San-sibar × Oase cross cultivated in Europe
3.1 Abstract 19
3.1 Abstract
Phytosterols are one of the minor seed constituents in oilseed rape that have received wide-interest in the food and nutrition industry due to its health benefit in lowering LDL cholesterol in humans.
To understand the genetic basis of phytosterol content and its relationship with other seed quality traits in oilseed rape, quantitative trait loci (QTL) mapping was performed in a segregating double-haploid (DH) population derived from the cross of two winter oilseed rape varieties "Sansibar"
and "Oase", termed SODH population. Both parental lines are of canola quality and were chosen due to their contrasting phytosterol and oil contents in seed. A genetic map was constructed for SODH population based on a total of 1642 markers (AFLP, candidate-gene based marker, DArT, Silico-DArT, SSR, and SNP), organized in 23 linkage groups and covering a map length of 2350 cM with a mean marker interval of 2.0 cM. The SODH population was cultivated at six environments in Europe and was phenotyped for phytosterol contents as well as some important seed quality traits such as oil content, fatty acid composition and protein content of defatted meal, and a yield related trait, seed weight. Multiple interval mapping identified 29 QTL for nine phytosterol traits, 16 QTL for four fatty acids, six QTL for oil content, four QTL for protein content of defatted meal and three QTL for seed weight. Colocalizations of QTL for different traits were more frequently observed than individual isolated QTL. Four genomic regions with major QTL (R2≥25%) were found for brassicasterol on A04, campesterol:sitosterol and 24-methyl:24-ethyl sterol on A06, C18:1 and C18:3 on A01, and C16:0 on A09. Possible candidate genes that underlie these four QTL genomic regions were revealed by aligning locations of QTL with the reference sequence ofBrassica rapa. A relatively good collinearity between genetic and physical map positions were observed in all four QTL genomic regions. QTL for brassicasterol on A04 was colocalized withCYP710A1, a gene that encodes the cytochrome P450 enzyme which might be responsible for converting 24-epi-campesterol to brassicasterol. QTL for campesterol:sitosterol and 24-methyl:24-ethyl sterol on A06 were colocalized withSMT2, a gene that encodes the sterol C24-methyltransferase 2 which converts 24-methylenelophenol to 24-ethylidene lophenol. QTL for C18:1 and C18:3 on A01 were colocalized withFAD2, a gene that encodes the endoplasmic delta-12 oleate desaturase which desaturate C18:1 into C18:2. QTL for C16:0 on A09 was colocalized withFATB, a gene that encodes the acyl-ACP thioesterase which hydrolyzes the thioester bond of C16:0-ACP and releases C16:0
and ACP.