Patterns of molecular variation in a species-wide germplasm set of <Emphasis Type="Italic">Brassica napus</Emphasis>

Rapeseed (Brassica napus L.) is the leading European oilseed crop serving as source for edible oil and renewable energy. The objectives of our study were to (i) examine the population structure of a large and diverse set of B. napus inbred lines, (ii) investigate patterns of genetic diversity within and among different germplasm types, (iii) compare the two genomes of B. napus with regard to genetic diversity, and (iv) assess the extent of linkage disequilibrium (LD) between simple sequence repeat (SSR) markers. Our study was based on 509 B. napus inbred lines genotyped with 89 genome-specific SSR primer combinations. Both a principal coordinate analysis and software STRUCTURE revealed that winter types, spring types, and swedes were assigned to three major clusters. The genetic diversity of winter oilseed rape was lower than the diversity found in other germplasm types. Within winter oilseed rape types, a decay of genetic diversity with more recent release dates and reduced levels of erucic acid and glucosinolates was observed. The percentage of linked SSR loci pairs in significant (r ² > Q _{95 unlinked loci pairs}) LD was 6.29% for the entire germplasm set. Furthermore, LD decayed rapidly with distance, which will allow a relatively high mapping resolution in genome-wide association studies using our germplasm set, but, on the other hand, will require a high number of markers.     

Variation in the glucosinolate content of vegetative tissues of Chinese lines of Brassica napus L

The glucosinolate content of leaves, stems and roots of a range of Chinese oilseed rape (Brassica napus L.) breeding lines was analysed. Total content and spectrum of individual glucosinolates varied widely, and there was no correlation between seed and vegetative tissue glucosinolate content. Lines with low seed glucosinolates (00) did not necessarily have low glucosinolate content in vegetative tissues; nor did high seed glucosinolate lines always have high vegetative tissue content. There was no correlation between the glucosinolate content of leaf, stem, and root in any given line. It appears that glucosinolate synthesis and accumulation is under tissue-specific control, and the mutation which blocks accumulation of glucosinolates in seeds does not influence other tissues. The responses of these lines to elicitors was also examined. Methyl jasmonate and salicylic acid treatments produced increases in leaf indolyl and aromatic glucosinolates respectively. However, the extent of such increases differed widely between the lines, and there were other, less consistent, effects on other classes of glucosinolate. There seems to be greater variation in glucosinolate accumulation in rape than has previously been reported, and the lines described here have considerable potential for evaluating the effects of manipulating glucosinolate profiles on pest and disease interactions.     

The identification and mapping of candidate genes and QTL involved in the fatty acid desaturation pathway in <Emphasis Type="Italic">Brassica napus</Emphasis>

We constructed a linkage map for the population QDH, which was derived from a cross between an oilseed rape cultivar and a resynthesised Brassica napus. The linkage map included ten markers linked to loci orthologous to those encoding fatty acid biosynthesis genes in Arabidopsis thaliana. The QDH population contains a high level of allelic variation, particularly in the C genome. We conducted quantitative trait locus (QTL) analyses, using field data obtained over 3 years, for the fatty acid composition of seed oil. The population segregates for the two major loci controlling erucic acid content, on linkage groups A8 and C3, which quantitatively affect the content of other fatty acids and is a problem generally encountered when crossing “wild” germplasm with cultivated “double low” oilseed rape cultivars. We assessed three methods for QTL analysis, interval mapping, multiple QTL mapping and single marker regression analysis of the subset of lines with low erucic acid. We found the third of these methods to be most appropriate for our main purpose, which was the study of the genetic control of the desaturation of 18-carbon fatty acids. This method enabled us to decouple the effect of the segregation of the erucic acid-controlling loci and identify 34 QTL for fatty acid content of seed oil, 14 in the A genome and 20 in the C genome. The QTL indicate the presence of 13 loci with novel alleles inherited from the progenitors of the resynthesised B. napus that might be useful for modulating the content or extent of desaturation of polyunsaturated fatty acids, only one of which coincides with the anticipated position of a candidate gene, an orthologue of FAD2.     

Transfer and segregation of triazine tolerant chloroplasts in Brassica napus L.

Summary Hypocotyl protoplasts of 45 different genotypes of German winter oilseed rape Brassica napus L. (double zero quality: high in yield, seeds low in erucic acid and glucosinolate content) were regenerated to plants. Triazine/triazinone (tri)-tolerant chloroplasts of the Canadian spring oilseed rape variety OAC Triton were introduced into some winter oilseed rapes by means of protoplast fusion. X-ray irradiation was used to limit the transfer of nuclear DNA of Triton protoplasts and to promote the selective transfer of tri-tolerant chloroplasts. Regenerated cybrid plants survived a treatment rate of 1000 g/ha metribuzin. The presence and segregation of the tri-tolerant chloroplasts in winter oilseed rape plants, regenerated from fusion products and their progeny, was investigated by restriction fragment length polymorphism (RFLP). Our results indicate that chloroplast segregation was not completed in plants regnerated from fusion products derived from X-irradiated OAC Triton mesophyll protoplasts and German winter oilseed rape hypocotyl protoplasts. In regenerants and their progeny both chloroplast types can still be present. Chloroplasts derived from wintertype protoplasts can outcompete tritolerant chloroplasts during plant development. In some instances, even progeny plants not kept under selective conditions (metribuzin) lost tri-tolerant chloroplasts. A homogenous population of tri-tolerant chloroplasts was necessary to obtain stable tri-tolerant winter oilseed rape plants.     

Genetic analysis of phenylpropanoid metabolites associated with resistance against Verticillium longisporum in Brassica napus

Verticillium longisporum is a major threat to production of oilseed rape (Brassica napus) in Europe. The aim of the study was to develop new markers and obtain insights into putative mechanisms and pathways involved in the resistance reaction. A genetic approach was used to identify quantitative trait loci (QTL) for V. longisporum resistance and metabolic traits potentially influencing resistance in a B. napus mapping population. Resistance to V. longisporum was mapped in a doubled haploid (DH) population from a cross between the partially resistant winter oilseed rape variety Express 617 and a resistant resynthesized B. napus line, R53. One major resistance QTL contributed by R53 was identified on chromosome C5, while a further, minor QTL contributed by Express 617 was detected on chromosome C1. Markers flanking the QTL also significantly correlated with V. longisporum resistance in four further DH populations derived from crosses between elite oilseed rape cultivars and other resynthesized B. napus lines originating from genetically and geographically diverse brassica A and C genome donors. The tightly-linked markers developed enable the combination of favorable alleles for novel resistance loci from resynthesized B. napus materials with existing resistance loci from commercial breeding lines. HPLC analysis of hypocotyls from infected DH lines revealed that concentrations of a number of phenylpropanoids were correlated with V. longisporum resistance. QTL for some of these phenylpropanoids were also found to co-localize with the QTL for V. longisporum resistance. Genes from the phenylpropanoid pathway are suggested as candidates for V. longisporum resistance.     

Constitutive and herbivore-inducible glucosinolate concentrations in oilseed rape (Brassica napus) leaves are not affected by Bt Cry1Ac insertion but change under elevated atmospheric CO2 and O3

Glucosinolates are plant secondary compounds involved in direct chemical defence by cruciferous plants against herbivores. The glucosinolate profile can be affected by abiotic and biotic environmental stimuli. We studied changes in glucosinolate patterns in leaves of non-transgenic oilseed rape (Brassica napus ssp. oleifera) under elevated atmospheric CO₂ or ozone (O₃) concentrations and compared them with those from transgenic for herbivore-resistance (Bacillus thuringiensis Cry1Ac endotoxin), to assess herbivory dynamics. Both elevated CO₂ and O₃ levels decreased indolic glucosinolate concentrations in transgenic and non-transgenic lines, whereas O₃ specifically increased the concentration of an aromatic glucosinolate, 2-phenylethylglucosinolate. The herbivore-inducible indolic glucosinolate response was reduced in elevated O₃ whereas elevated CO₂ altered the induction dynamics of indolic and aliphatic glucosinolates. Herbivore-resistant Bt plants experienced minimal leaf damage after target herbivore Plutella xylostella feeding, but exhibited comparatively similar increase in glucosinolate concentrations after herbivory as non-transgenic plants, indicating that the endogenous glucosinolate defence was not severely compromised by transgenic modifications. The observed differences in constitutive and inducible glucosinolate concentrations of oilseed rape under elevated atmospheric CO₂ and O₃ might have implications for plant–herbivore interactions in Brassica crop-ecosystems in future climate scenarios.     

Quantitative trait analysis of seed yield and other complex traits in hybrid spring rapeseed (Brassica napus L.): 1. Identification of genomic regions from winter germplasm

The introgression of winter germplasm into spring canola (Brassica napus L.) represents a novel approach to improve seed yield of hybrid spring canola. In this study, quantitative trait loci (QTL) for seed yield and other traits were genetically mapped to determine the effects of genomic regions introgressed from winter germplasm into spring canola. Plant materials used comprised of two populations of doubled haploid (DH) lines having winter germplasm introgression from two related French winter cultivars and their testcrosses with a spring line used in commercial hybrids. These populations were evaluated for 2 years at two locations (Wisconsin, USA and Saskatchewan, Canada). Genetic linkage maps based on RFLP loci were constructed for each DH population. Six QTL were detected in the testcross populations for which the winter alleles increased seed yield. One of these QTL explained 11 and 19% of the phenotypic variation in the two Canadian environments. The winter allele for another QTL that increased seed yield was linked in coupling to a QTL allele for high glucosinolate content, suggesting that the transition of rapeseed into canola could have resulted in the loss of favorable seed yield alleles. Most QTL for which the introgressed allele decreased seed yield of hybrids mapped to genomic regions having homoeologous non-reciprocal transpositions. This suggests that allelic configurations created by these rearrangements might make an important contribution to genetic variation for complex traits in oilseed B. napus and could account for a portion of the heterotic effects in hybrids. Electronic Supplementary Material Supplementary material is available to authorised users in the online version of this article at .     

The effect of modifying the glucosinolate content of leaves of oilseed rape (Brassica napus ssp. oleifera) on its interaction with specialist and generalist pests

Twenty eight Brassica napus lines were developed which had contrasting leaf glucosinolate profiles to those found in commercial oilseed rape cultivars. The lines varied both in the total amount of aliphatic glucosinolates and in the ratio of different side chain structures. The lines were used in field experiments to assess the manner by which glucosinolates mediate the interactions between Brasssica and specialist pests (Psylliodes chrysocephala and Pieris rapae) and generalist pests (pigeons and slugs). Increases in the level of glucosinolates resulted in greater damage by adult flea beetles (P. chrysocephala) and a greater incidence of Pieris rapae larvae, but reduced the extent of grazing by pigeons and slugs. Decreasing the side chain length of aliphatic glucosinolates and reducing the extent of hydroxylation of butenyl glucosinolates increased the extent of adult flea beetle feeding. The implications of modifying the glucosinolate content of the leaves of oilseed rape and the role of these secondary metabolites in plant/herbivore interactions are discussed.     

Quantitative trait analysis of seed yield and other complex traits in hybrid spring rapeseed (Brassica napus L.): 2. Identification of alleles from unadapted germplasm

Unadapted germplasm may contain alleles that could improve hybrid cultivars of spring oilseed Brassica napus. Quantitative trait loci (QTL) mapping was used to identify potentially useful alleles from two unadapted germplasm sources, a Chinese winter cultivar and a re-synthesized B. napus, that increase seed yield when introgressed into a B. napus spring hybrid combination. Two populations of 160 doubled haploid (DH) lines were created from crosses between the unadapted germplasm source and a genetically engineered male-fertility restorer line (P1804). A genetically engineered male-sterile tester line was used to create hybrids with each DH line (testcrosses). The two DH line populations were evaluated in two environments and the two testcross populations were evaluated in three or four environments for seed yield and other agronomic traits. Several genomic regions were found in the two testcross populations which contained QTL for seed yield. The map positions of QTL for days to flowering and resistance to a bacterial leaf blight disease coincided with QTL for seed yield and other agronomic traits, suggesting the occurrence of pleiotropic or linked effects. For two hybrid seed yield QTL, the favorable alleles increasing seed yield originated from the unadapted parents, and one of these QTL was detected in multiple environments and in both populations. In this QTL region, a chromosome rearrangement was identified in P1804, which may have affected seed yield.Electronic Supplementary Material Supplementary material is available to authorised users in the online version of this article at .     

Genetic diversity of oilseedBrassica napus germ plasm based on restriction fragment length polymorphisms

Oilseed rape (Brassica napus) is an important oilseed crop worldwide. Cultivars have been developed for many growing regions, however little is known about genetic diversity inB. napus germ plasm. The purpose of the research presented here was to study the genetic diversity and relationships ofB. napus accessions using restriction fragment length polymorphisms (RFLPs). Eighty threeB. napus accessions were screened using 43 genomic DNA clones which revealed 161 polymorphic fragments. Each accession was uniquely identified by the markers with the exception of the near-isogenic cvs Triton and Tower. The RFLP data were analyzed by cluster analysis of similarity coefficients and by principal component analysis. Overall, there were three major groups of cultivars. The first group included only spring accessions, the second mostly winter accessions and the third, rutabagas and oilseed rape accessions from China and Japan. These results indicate that withinB. napus, winter and spring cultivars represent genetically distinct groups. The grouping of accessions by cluster analysis was generally consistent with known pedigrees. This consistency included the grouping of lines derived both by backcrossing or self-pollination with their parents.     

Analysis of QTLs for emcic acid and oil content in seeds on A8 chro- mosome and the linkage drag between the alleles for the two traits in Brassica napus

The history of canola breeding began with the discovery of germplasm with low erucic acid content in seeds of spring forage cultivar in the 1950's. FAE1 mutations led to a dramatic decrease of the seed erucic acid content in Arabidopsis thaliana. The products of the two FAE1 loci, BnA8.FAE1 and BnC3.FAE1, showed additive effects to the level of erucic acid content in oilseed rape. Previous research believed that the pleiotropy of FAE1 was responsible for the decrease in seed oil content along with the reduction of seed erucic acid content in the modern cultivars. TN DH population was developed from a canola cultivar Tapidor and a Chinese traditional cultivar Ningyou7. The population had been tested in 10 and 11 environments to map QTLs for the erucic acid content and oil content in seeds. As the map resolution increased, a novel QTL for seed erucic acid content was revealed, after Meta-analysis, 7 cM away from the most significant seed erucic acid content QTL where BnA8.FAE1 is located. Seven independent QTLs for seed oil content (qOC) were detected around the two seed erucic acid content QTLs (qEA) across 39.20 cM on linkage group A8. Two of the qOCs co-localized with the two qEAs, respectively, and were detected in a single environment. The other five qOCs were detected in 10 of 11 environments independent of qEAs. Alleles from Tapidor in all the QTLs at the 0–39.20 cM region contributed negative effects to either erucic acid content or oil content in seeds. Parallel, genotyping showed that on 5 of the 7 QTLs regions, Tapidor alleles had the same genotypes with that in ‘Liho’, the original low seed erucic acid content source. Through rounds of crossbreeding with oil-cropped cultivars and intensive selection for multi generations, Tapidor still had the inferior alleles for low seed oil content from ‘Liho’, the forage rape. This showed a strong linkage drag of low seed oil content, which was controlled by the five qEA-independent qOCs, with low seed erucic acid content. Ninety cultivars of B. napus from 8 countries were used to analyze the genetic drag with 9 molecular markers located in the QTL confidence intervals (24.04 cM) on linkage group A8. It was noticed that more than 46% of the cultivars with low seed erucic acid content trait remained the genotype of low seed oil content at least in one locus. Backcross and marker-assisted selection could break the genetic drag between the low oil content and erucic acid in seeds in the process for breeding modern high seed oil content canola cultivars.     

RFLP mapping of quantitative trait loci controlling seed aliphatic-glucosinolate content in oilseed rape (Brassica napus L)

We report the RFLP mapping of quantitative trait loci (QTLs) which regulate the total seed aliphaticglucosinolate content in Brassica napus L. A population of 99 F₁-derived doubled-haploid (DH) recombinant lines from a cross between the cultivars Stellar (low-glucosinolate) and Major (high-glucosinolate) was used for singlemarker analysis and the interval mapping of QTLs associated with total seed glucosinolates. Two major loci, GSL-1 and GSL-2, with the largest influence on total seed aliphatic-glucosinolates, were mapped onto LG 20 and LG 1, respectively. Three loci with smaller effects, GSL-3, GSL-4 and GSL-5, were tentatively mapped to LG 18, LG 4 and LG 13, respectively. The QTLs acted in an additive manner and accounted for 71 % of the variation in total seed glucosinolates, with GSL-1 and GSL-2 accounting for 33% and 17%, respectively. The recombinant population had aliphatic-glucosinolate levels of between 6 and 160 moles per g^-1 dry wt of seed. Transgressive segregation for high seed glucosinolate content was apparent in 25 individuals. These phenotypes possessed Stellar alleles at GSL-3 and Major alleles at the four other GSL loci demonstrating that low-glucosinolate genotypes (i.e. Stellar) may possess alleles for high glucosinolates which are only expressed in particular genetic backgrounds. Gsl-elong and Gsl-alk, loci which regulate the ratio of individual aliphatic glucosinolates, were also mapped. Gsl-elong-1 and Gsl-elong-2, which control elongation of the -amino-acid precursors, mapped to LG 18 and LG 20 and were coincident with GSL loci which regulate total seed aliphatic glucosinolates. A third tentative QTL, which regulates side-chain elongation, was tentatively mapped to LG 12. Gsl-alk, which regulates H₃CS-removal and side-chain de-saturation, mapped to LG 20.     

Effects of Glucosinolates and Flavonoids on Colonization of the Roots of Brassica napus by Azorhizobium caulinodans ORS571

Plants of Brassica napus were assessed quantitatively for their susceptibility to lateral root crack colonization by Azorhizobium caulinodans ORS571(pXLGD4) (a rhizobial strain carrying the lacZ reporter gene) and for the concentration of glucosinolates in their roots by high-pressure liquid chromatography (HPLC). High- and low-glucosinolate-seed (HGS and LGS) varieties exhibited a relatively low and high percentage of colonized lateral roots, respectively. HPLC showed that roots of HGS plants contained a higher concentration of glucosinolates than roots of LGS plants. One LGS variety showing fewer colonized lateral roots than other LGS varieties contained a higher concentration of glucosinolates than other LGS plants. Inoculated HGS plants treated with the flavonoid naringenin showed significantly more colonization than untreated HGS plants. This increase was not mediated by a naringenin-induced lowering of the glucosinolate content of HGS plant roots, nor did naringenin induce bacterial resistance to glucosinolates or increase the growth of bacteria. The erucic acid content of seed did not appear to influence colonization by azorhizobia. Frequently, leaf assays are used to study glucosinolates and plant defense; this study provides data on glucosinolates and bacterial colonization in roots and describes a bacterial reporter gene assay tailored easily to the study of ecologically important phytochemicals that influence bacterial colonization. These data also form a basis for future assessments of the benefits to oilseed rape plants of interaction with plant growth-promoting bacteria, especially diazotrophic bacteria potentially able to extend the benefits of nitrogen fixation to nonlegumes.     

Mapping of quantitative trait loci and development of allele-specific markers for seed weight in Brassica napus

Seed weight is an important component of grain yield in oilseed rape (Brassica napus L.), but the genetic basis for the important quantitative trait is still not clear. In order to identify the genes for seed weight in oilseed rape, QTL mapping for thousand seed weight (TSW) was conducted with a doubled haploid (DH) population and an F₂ population. A complete linkage map of the DH population was constructed using 297 simple sequence repeat (SSR) markers. Among nine TSW QTLs detected, two major QTLs, TSWA7a and TSWA7b, were stably identified across years and collectively explained 27.6–37.9% of the trait variation in the DH population. No significant epistatic interactions for TSW detected in the DH population indicate that the seed weight variation may be primarily attributed to additive effects. The stability and significance of TSWA7a and TSWA7b were further validated in the F₂ population with different genetic backgrounds. By cloning BnMINI3a and BnTTG2a, two B. napus homologous genes to Arabidopsis thaliana, allele-specific markers were developed for TSWA5b and TSWA5c, two TSW QTLs on A5, respectively. The importance of the major and minor QTLs identified was further demonstrated by analysis of the allelic effects on TSW in the DH population.     

A comparative linkage map of oilseed rape and its use for QTL analysis of seed oil and erucic acid content

We have developed a new DH mapping population for oilseed rape, named TNDH, using genetically and phenotypically diverse parental lines. We used the population in the construction of a high stringency genetic linkage map, consisting of 277 loci, for use in quantitative genetic analysis. A proportion of the markers had been used previously in the construction of linkage maps for Brassica species, thus permitting the alignment of maps. The map includes 68 newly developed Sequence Tagged Site (STS) markers targeted to the homologues of defined genes of A. thaliana. The use of these markers permits the alignment of our linkage map with the A. thaliana genome sequence. An additional 74 loci (31 newly developed STS markers and 43 loci defined by SSR and RFLP markers that had previously been used in published linkage maps) were added to the map. These markers increased the resolution of alignment of the newly constructed linkage map with existing Brassica linkage maps and the A. thaliana genome sequence. We conducted field trials with the TNDH population at two sites, and over 2 years, and identified reproducible QTL for seed oil content and erucic acid content. The results provide new insights into the genetic control of seed oil and erucic acid content in oilseed rape, and demonstrate the utility of the linkage map and population.Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.D. Qiu and C. Morgan authors contributed equally to the work.     

The influence of glucosinolates and sugars on feeding by the cabbage stem flea beetle,Psylliodes chrysocephala

Agar was used as an artificial substrate to investigate the feeding behaviour of the cabbage stem flea beetle,Psylliodes chrysocephala L. (Coleoptera: Chrysomelidae), an important pest of winter oilseed rape (Brassica napus) in Europe. Both glucosinolates and sugars stimulated feeding when added to agar. The amount of feeding that occurred was affected by the type and concentration of glucosinolate and surgar and also by combinations of components. Although glucosinolates were potent feeding stimulants forP. chrysocephala, they were not a prerequisite for feeding, nor does it seem likely that glucosinolate profiles are used by this species to discriminate amongst cruciferous plants at the gustatory level.     

Genetics of aliphatic glucosinolates. IV. Side-chain modification in Brassica oleracea

The biochemical and genetical relationship between aliphatic glucosinolates which have methylthioalkyl, methylsulphinylalkyl and alkenyl side chains has not been resolved by biochemical studies. In this study, two hypothetical models are tested by the genetic analysis of a backcross population between Brassica drepanensis and B. atlantica. The results support one of the models in which 3-methylthiopropyl glucosinolate is sequentially converted to 3-methylsulphinylpropyl, and then to 2-propenyl glucosinolate, by the action of dominant alleles at two loci. RFLP mapping positioned both loci on the same linkage group homologous to the B. napus N19 linkage group. The implication of the results for the genetic manipulation of glucosinolates in Brassica to improve flavour and nutritional properties, and in order to investigate plant-insect interactions, is discussed.     

Developmental regulation of aldoxime formation in seedlings and mature plants of Chinese cabbage (Brassica campestris ssp. pekinensis) and oilseed rape (Brassica napus): Glucosinolate and IAA biosynthetic enzymes

The first steps in the biosynthesis of glucosinolates and indole-3-acetic acid (IAA) in oilseed rape (Brassica napus L.) and Chinese cabbage (Brassica campestris ssp. pekinensis) involve the formation of aldoximes. In rape the formation of aldoximes from chain-extended amino acids, for aromatic and aliphatic glucosinolate biosynthesis, is catalysed by microsomal flavin-containing monooxygenases. The formation of indole-3-aldoxime from l-tryptophan, the potential precursor of both indole-3-acetic acid and indolyl-glucosinolates, is catalysed by several microsomal peroxidases. The biosynthesis of glucosinolates and indole-3-acetic acid was shown to be under developmental control in oilseed rape and Chinese cabbage. No monooxygenase activities were detected in cotyledons or old leaves of either species. The highest monooxygenase activities were found in young expanding leaves; as the leaves reached full expansion and matured the activities decreased rapidly. The indole-aldoxime-forming activity was found in all of the tissues analysed, but there was also a clear decrease in foliar activity with maturity in leaves of rape and Chinese cabbage. Partial characterisation of the Chinese cabbage monooxygenases showed that they have essentially identical properties to the previously characterised rape enzymes; they are not cytochrome P450-type enzymes, but resemble flavin-containing monooxygenases. No monooxygenase inhibitors were detected in microsomes prepared from either cotyledons or old leaves.Abbreviations DHMet dihomomethionine - FMO flavin-containing monooxygenase - HPhe homophenylalanine - IAA indole-3-acetic acid - l-Phe l-phenylalanine - l-Trp l-tryptophan - MO monooxygenase - IAALD indole-3-acetaldehyde - IAOX indole-3-aldoxime - THMet trihomomethionine     

Genomic microstructure and differential expression of the genes encoding UDP-glucose:sinapate glucosyltransferase (UGT84A9) in oilseed rape (Brassica napus)

In oilseed rape (Brassica napus), the glucosyltransferase UGT84A9 catalyzes the formation of 1-O-sinapoyl-β-glucose, which feeds as acyl donor into a broad range of accumulating sinapate esters, including the major antinutritive seed component sinapoylcholine (sinapine). Since down-regulation of UGT84A9 was highly efficient in decreasing the sinapate ester content, the genes encoding this enzyme were considered as potential targets for molecular breeding of low sinapine oilseed rape. B. napus harbors two distinguishable sequence types of the UGT84A9 gene designated as UGT84A9-1 and UGT84A9-2. UGT84A9-1 is the predominantly expressed variant, which is significantly up-regulated during the seed filling phase, when sinapate ester biosynthesis exhibits strongest activity. In the allotetraploid genome of B. napus, UGT84A9-1 is represented by two loci, one derived from the Brassica C-genome (UGT84A9a) and one from the Brassica A-genome (UGT84A9b). Likewise, for UGT84A9-2 two loci were identified in B. napus originating from both diploid ancestor genomes (UGT84A9c, Brassica C-genome; UGT84A9d, Brassica A-genome). The distinct UGT84A9 loci were genetically mapped to linkage groups N15 (UGT84A9a), N05 (UGT84A9b), N11 (UGT84A9c) and N01 (UGT84A9d). All four UGT84A9 genomic loci from B. napus display a remarkably low micro-collinearity with the homologous genomic region of Arabidopsis thaliana chromosome III, but exhibit a high density of transposon-derived sequence elements. Expression patterns indicate that the orthologous genes UGT84A9a and UGT84A9b should be considered for mutagenesis inactivation to introduce the low sinapine trait into oilseed rape.