Maoteng Li

Introgression of genomic components from Chinese Brassica rapa contributes to widening the genetic diversity in rapeseed (B. napus L.), with emphasis on the evolution of Chinese rapeseed

In spite of its short history of being an oil crop in China, the Chinese semi-winter rapeseed (Brassica napus L., 2nxa0=xa038, AACC) has been improved rapidly by intentional introgression of genomic components from Chinese B. rapa (2nxa0=xa020, AA). As a result, the Chinese semi-winter rapeseed has diversified genetically from the spring and winter rapeseed grown in the other regions such as Europe and North America. The objectives of this study were to investigate the roles of the introgression of the genomic components from the Chinese B. rapa in widening the genetic diversity of rapeseed and to verify the role of this introgression in the evolution of the Chinese rapeseed. Ten lines of the new type of rapeseed, which were produced by introgression of Chinese B. rapa to Chinese normal rapeseed, were compared for genetic diversity using amplified fragment length polymorphism (AFLP) with three groups of 35 lines of the normal rapeseed, including 9 semi-winter rapeseed lines from China, 9 winter rapeseed lines from Europe and 17 spring rapeseed lines from Northern Europe, Canada and Australia. Analysis of 799 polymorphic fragments revealed that within the groups, the new type rapeseed had the highest genetic diversity, followed by the semi-winter normal rapeseed from China. Spring and winter rapeseed had the lowest genetic diversity. Among the groups, the new type rapeseed group had the largest average genetic distance to the other three groups. Principal component analysis and cluster analysis, however, could not separate the new type rapeseed group from Chinese normal rapeseed group. Our data suggested that the introgression of Chinese B. rapa could significantly diversify the genetic basis of the rapeseed and play an important role in the evolution of Chinese rapeseed. The use of new genetic variation for the exploitation of heterosis in Brassica hybrid breeding is discussed

Heterotic patterns in rapeseed (Brassica napus L.): I. Crosses between spring and Chinese semi-winter lines

Chinese semi-winter rapeseed is genetically diverse from Canadian and European spring rapeseed. This study was conducted to evaluate the potential of semi-winter rapeseed for spring rapeseed hybrid breeding, to assess the genetic effects involved, and to estimate the correlation of parental genetic distance (GD) with hybrid performance, heterosis, general combining ability (GCA) and specific combining ability (SCA) in crosses between spring and semi-winter rapeseed lines. Four spring male sterile lines from Germany and Canada as testers were crossed with 13 Chinese semi-winter rapeseed lines to develop 52 hybrids, which were evaluated together with their parents and commercial hybrids for seed yield and oil content in three sets of field trials with 8 environments in Canada and Europe. The Chinese parental lines were not adapted to local environmental conditions as demonstrated by poor seed yields per se. However, the hybrids between the Chinese parents and the adapted spring rapeseed lines exhibited high heterosis for seed yield. The average mid-parent heterosis was 15% and ca. 50% of the hybrids were superior to the respective hybrid control across three sets of field trials. Additive gene effects mainly contributed to hybrid performance since the mean squares of GCA were higher as compared to SCA. The correlation between parental GD and hybrid performance and heterosis was found to be low whereas the correlation between GCA(fxa0+xa0m) and hybrid performance was high and significant in each set of field trials, with an average of rxa0=xa00.87 for seed yield and rxa0=xa00.89 for oil content, indicating that hybrid performance can be predicted by GCA(fxa0+xa0m). These results demonstrate that Chinese semi-winter rapeseed germplasm has a great potential to increase seed yield in spring rapeseed hybrid breeding programs in Canada and Europe.

Identification of QTLs Associated with Oil Content in a High-Oil Brassica napus Cultivar and Construction of a High-Density Consensus Map for QTLs Comparison in B. napus

Increasing seed oil content is one of the most important goals in breeding of rapeseed (B. napus L.). To dissect the genetic basis of oil content in B. napus, a large and new double haploid (DH) population containing 348 lines was obtained from a cross between ‘KenC-8’ and ‘N53-2’, two varieties with >10% difference in seed oil content, and this population was named the KN DH population. A genetic linkage map consisting of 403 markers was constructed, which covered a total length of 1783.9 cM with an average marker interval of 4.4 cM. The KN DH population was phenotyped in eight natural environments and subjected to quantitative trait loci (QTL) analysis for oil content. A total of 63 identified QTLs explaining 2.64–17.88% of the phenotypic variation were identified, and these QTLs were further integrated into 24 consensus QTLs located on 11 chromosomes using meta-analysis. A high-density consensus map with 1335 marker loci was constructed by combining the KN DH map with seven other published maps based on the common markers. Of the 24 consensus QTLs in the KN DH population, 14 were new QTLs including five new QTLs in A genome and nine in C genome. The analysis revealed that a larger population with significant differences in oil content gave a higher power detecting new QTLs for oil content, and the construction of the consensus map provided a new clue for comparing the QTLs detected in different populations. These findings enriched our knowledge of QTLs for oil content and should be a potential in marker-assisted breeding of B. napus.

Improvement of arachidonic acid and eicosapentaenoic acid production by increasing the copy number of the genes encoding fatty acid desaturase and elongase into Pichia pastoris.

Genes encoding Δ6 desaturase, Δ6 fatty acid elongase, and Δ5 desaturase from the alga, Phaeodactylum tricornutum, were co-expressed in Pichia pastoris to produce arachidonic acid (ARA; 20:4 Δ5,xa08,xa011,xa014) and eicosapentaenoic acid (EPA; 20:5 Δ5,xa08,xa011,xa014,xa017). A panel of Pichia clones carrying progressively increasing copies of the heterologous gene expression cassette was created using an in vitro multimerization approach. ARA and EPA accumulated up to 0.3 and 0.1% of total fatty acids, respectively, in the recombinant P. pastoris carrying with double copies of these three heterologous genes, as compared to 0.1 and 0.05%, respectively, in the recombinant P. pastoris with single copy.

Gene expression profiles associated with intersubgenomic heterosis in Brassica napus

In order to understand the genetic mechanism of heterosis that has been observed in hybrids between Brassica napus and partial new-type B. napus which had exotic genome components from relative species, this study focused on the difference in gene expression patterns among partial new-typed B. napus lines, B. napus cultivars and their hybrids using the cDNA amplified fragment length polymorphism technique (cDNA-AFLP) technique. First, three partial new-type B. napus lines were compared with their original parents. One new line contained the exotic genomic components from B. rapa, and the other two new lines were obtained by the introgression of genomic components from B. rapa and B. carinata. The experimental results showed that the introgression of Ar and Cc genome components from B. rapa and B. carinata led to considerable differences in the gene expression profiles of the partial new-type lines when compared with their parents. Secondly, the gene expression profiles of nine cross-combinations between three partial new-type lines and three B. napus cultivars were compared. Twenty transcript-derived fragments (TDFs) associated with intersubgenomic heterosis were randomly selected and converted into PCR-based molecular markers. Some of them were mapped in the confidence intervals of quantitative trait loci (QTLs) for yield and yield-related traits in three segregative populations of B. napus. These results suggested that a proportion of the heterosis-associated TDFs were really responsible for fluctuating seed yield in rapeseed.

Dynamic and comparative QTL analysis for plant height in different developmental stages of Brassica napus L.

AbstractKey messageThis report describes a dynamic QTL analysis for plant height at various stages using a large doubled haploid population and performs a QTL comparison between different populations inBrassica napus.AbstractPlant height (PH) not only plays an important role in determining plant architecture, but is also an important character related to yield. The process of determining PH occurs through a series of steps; however, no studies have focused on developmental behavior factors affecting PH in Brassica napus. In the present study, KN DH, a large doubled haploid population containing 348 lines was used for a dynamic quantitative trait locus (QTL) analysis for PH in six experiments. In all, 20 QTLs were identified at maturity, whereas 50 QTLs were detected by conditional mnapping method and the same number was identified by unconditional mapping strategies. Interestingly, five unconditional QTLs ucPH.A2-2, ucPH.A3-2, ucPH.C5-1, ucPH.C6-2 and ucPH.C6-3 were identified that were consistent over the all growth stages of one or two particular experiments, and one conditional QTL cPH.A2-3 was expressed throughout the entire growth process in one experiment. A total of 70 QTLs were obtained after combining QTLs identified at maturity, by conditional and unconditional mapping strategies, in which 25 showed opposite genetic effects in different periods/stages and experiments. A consensus map containing 1357 markers was constructed to compare QTLs identified in the KN population with five previously mapped populations. Alignment of the QTLs detected in different populations onto the consensus map showed that 27 were repeatedly detected in different genetic backgrounds. These findings will enhance our understanding of the genetic control of PH regulation in B. napus, and will be useful for rapeseed genetic manipulation through molecular marker-assisted selection.

Genome wide analysis of flowering time trait in multiple environments via high-throughput genotyping technique in Brassica napus L

The prediction of the flowering time (FT) trait in Brassica napus based on genome-wide markers and the detection of underlying genetic factors is important not only for oilseed producers around the world but also for the other crop industry in the rotation system in China. In previous studies the low density and mixture of biomarkers used obstructed genomic selection in B. napus and comprehensive mapping of FT related loci. In this study, a high-density genome-wide SNP set was genotyped from a double-haploid population of B. napus. We first performed genomic prediction of FT traits in B. napus using SNPs across the genome under ten environments of three geographic regions via eight existing genomic predictive models. The results showed that all the models achieved comparably high accuracies, verifying the feasibility of genomic prediction in B. napus. Next, we performed a large-scale mapping of FT related loci among three regions, and found 437 associated SNPs, some of which represented known FT genes, such as AP1 and PHYE. The genes tagged by the associated SNPs were enriched in biological processes involved in the formation of flowers. Epistasis analysis showed that significant interactions were found between detected loci, even among some known FT related genes. All the results showed that our large scale and high-density genotype data are of great practical and scientific values for B. napus. To our best knowledge, this is the first evaluation of genomic selection models in B. napus based on a high-density SNP dataset and large-scale mapping of FT loci.

Cell death unlikely contributes to taxol production in fungal elicitor-induced cell suspension cultures of Taxus chinensis.

Cell suspension cultures ofTaxus chinensis, with 20, 40 and 100xa0mg fungal elicitorxa0l−1 from Aspergillus niger, underwent rapid cell death after 24 h, which was about 2, 3.7 and 5-fold of that of the control. At the same time, Taxol production was increased, respectively, to about 5, 8 and 3-fold of that of the control. Inhibition of phenolics biosynthesis resulted in a 150% increase in cell death but a 54% decrease in Taxol production compared with 40 mg elicitorxa0l−1 alone. O2-free N2 inhibited cell death but had little effect on Taxol production as induced by 40xa0mg fungal elicitorxa0l−1.

Δ6-Desaturase from Mortierella alpina: cDNA cloning, expression, and phylogenetic analysis

The open reading frame of the Δ6-desaturase gene was isolated from Mortierella alpina W15 and the gene was cloned into a pPIC3.5K vector. The vector was transformed into Pichia pastoris GS115 and expression was induced with methanol. The Δ6-desaturase expressed in P. pastoris GS115 catalyzed the conversion of linoleic acid to γ-linolenic acid but not the conversion of α-linolenic acid to octadecatetraenoic acid. The results indicate that the Δ6-desaturase gene from M. alpina W15 has substrate specificity in different organisms. Phylogenetic analysis revealed that Δ6-desaturase genes can be divided into four monophyletic groups. This work paves the way for further study of the functions of Δ6-desaturase in fatty acid metabolism and its three-dimensional structure.

Cytoplasmic and Genomic Effects on Non-Meiosis-Driven Genetic Changes in Brassica Hybrids and Allotetraploids from Pairwise Crosses of Three Cultivated Diploids

Nuclear-cytoplasmic interactions are predicted to be important in shaping the genetic changes in early stage of allopolyploidization. Our previous study shows the specific role of genome and cytoplasm affecting the chromosome pairing in Brassica hybrids and allotetraploids from pairwise crosses between three cultivated diploids with A, B and C genomes, respectively. Herein, to address how parental genomes and cytoplasm affects genomic, epigenetic and gene expression changes prior to meiosis in these hybrids and allopolyploids, their patterns of AFLP (Amplified fragment length polymorphism), mAFLP (Methylation AFLP) and cDNA-AFLP were compared with the progenitors, revealing the major absent bands within each genome. These changes varied under various cytoplasm backgrounds and genome combinations, following the significant order of AFLP> mAFLP> cDNA -AFLP. The frequencies of AFLP bands lost were positively correlated with the divergence degrees of parental genomes, but not obvious for those of mAFLP and cDNA-AFLP absent bands, and methylation change showed least variations among hybrids and within each genome. These changes within each genome followed the A>B>C hierarchy, except the highest rate of cDNA loss in B genome. Among three changes, only overall AFLP bands were significantly correlated with cDNA-AFLP, and their correlations varied within each genome. These changes in allotetraploids were mainly caused by genome merger rather than doubling. Parental genomes altered differently at three levels, responded to the types of cytoplasm and genome and their interaction or divergence. The result provides new clues for instant non-meiosis-driven genome restructuring following genome merger and duplication.