Analysing the genetic architecture of clubroot resistance variation in Brassica napus by associative transcriptomics

Clubroot is a destructive soil-borne pathogen of Brassicaceae that causes significant recurrent reductions in yield of cruciferous crops. Although there is some resistance in oilseed rape (a crop type of the species Brassica napus), the genetic basis of that resistance is poorly understood. In this study, we used an associative transcriptomics approach to elucidate the genetic basis of resistance to clubroot pathotype ECD 17/31/31 across a genetic diversity panel of 245 accessions of B. napus. A single nucleotide polymorphism (SNP) association analysis was performed with 256,397 SNPs distributed across the genome of B. napus and combined with transcript abundance data of 53,889 coding DNA sequence (CDS) gene models. The SNP association analysis identified two major loci (on chromosomes A2 and A3) controlling resistance and seven minor loci. Within these were a total of 86 SNP markers. Altogether, 392 genes were found in these regions. Another 21 genes were implicated as potentially involved in resistance using gene expression marker (GEM) analysis. After GO enrichment analysis and InterPro functional analysis of the identified genes, 82 candidate genes were identified as having roles in clubroot resistance. These results provide useful information for marker-assisted breeding which could lead to acceleration of pyramiding of multiple clubroot resistance genes in new varieties

Extensive homoeologous genome exchanges in allopolyploid crops revealed by mRNAseq-based visualization

Polyploidy, the possession of multiple sets of chromosomes, has been a predominant factor in the evolution and success of the angiosperms. Although artificially formed allopolyploids show a high rate of genome rearrangement, the genomes of cultivars and germplasm used for crop breeding were assumed stable and genome structural variation under the artificial selection process of commercial breeding has remained little studied. Here, we show, using a repurposed visualization method based on transcriptome sequence data, that genome structural rearrangement occurs frequently in varieties of three polyploid crops (oilseed rape, mustard rape and bread wheat), meaning that the extent of genome structural variation present in commercial crops is much higher than expected. Exchanges were found to occur most frequently where homoeologous chromosome segments are collinear to telomeres and in material produced as doubled haploids. The new insights into genome structural evolution enable us to reinterpret the results of recent studies and implicate homoeologous exchanges, not deletions, as being responsible for variation controlling important seed quality traits in rapeseed. Having begun to identify the extent of genome structural variation in polyploid crops, we can envisage new strategies for the global challenge of broadening crop genetic diversity and accelerating adaptation, such as the molecular identification and selection of genome deletions or duplications encompassing genes with trait-controlling dosage effects

A Brassica napus Reductase Gene Dissected by Associative Transcriptomics Enhances Plant Adaption to Freezing Stress

Cold treatment (vernalization) is required for winter crops such as rapeseed (Brassica napus L.). However, excessive exposure to low temperature (LT) in winter is also a stress for the semi-winter, early-flowering rapeseed varieties widely cultivated in China. Photosynthetic efficiency is one of the key determinants, and thus a good indicator for LT tolerance in plants. So far, the genetic basis underlying photosynthetic efficiency is poorly understood in rapeseed. Here the current study used Associative Transcriptomics to identify genetic loci controlling photosynthetic gas exchange parameters in a diversity panel comprising 123 accessions. A total of 201 significant Single Nucleotide Polymorphisms (SNPs) and 147 Gene Expression Markers (GEMs) were detected, leading to the identification of 22 candidate genes. Of these, Cab026133.1, an ortholog of the Arabidopsis gene AT2G29300.2 encoding a tropinone reductase (BnTR1), was further confirmed to be closely linked to transpiration rate. Ectopic expressing BnTR1 in Arabidopsis plants significantly increased the transpiration rate and enhanced LT tolerance under freezing conditions. Also, a much higher level of alkaloids content was observed in the transgenic Arabidopsis plants, which could help protect against LT stress. Together, the current study showed that AT is an effective approach for dissecting LT tolerance trait in rapeseed and that BnTR1 is a good target gene for the genetic improvement of LT tolerance in plant

Characterization of Chicken Spleen Transcriptome after Infection with <em>Salmonella enterica</em> Serovar Enteritidis

<div><p>In this study we were interested in identification of new markers of chicken response to <em>Salmonella</em> Enteritidis infection. To reach this aim, gene expression in the spleens of naive chickens and those intravenously infected with <em>S</em>. Enteritidis with or without previous oral vaccination was determined by 454 pyrosequencing of splenic mRNA/cDNA. Forty genes with increased expression at the level of transcription were identified. The most inducible genes encoded avidin (AVD), extracellular fatty acid binding protein (EXFABP), immune responsive gene 1 (IRG1), chemokine ah221 (AH221), trappin-6-like protein (TRAP6) and serum amyloid A (SAA). Using cDNA from sorted splenic B-lymphocytes, macrophages, CD4, CD8 and γδ T-lymphocytes, we found that the above mentioned genes were preferentially expressed in macrophages. AVD, EXFABP, IRG1, AH221, TRAP6 and SAA were induced also in the cecum of chickens orally infected with <em>S</em>. Enteritidis on day 1 of life or day 42 of life. Unusual results were obtained for the immunoglobulin encoding transcripts. Prior to the infection, transcripts coding for the constant parts of IgM, IgY, IgA and Ig light chain were detected in B-lymphocytes. However, after the infection, immunoglobulin encoding transcripts were expressed also by T-lymphocytes and macrophages. Expression of AVD, EXFABP, IRG1, AH221, TRAP6, SAA and all immunoglobulin genes can be therefore used for the characterization of the course of <em>S</em>. Enteritidis infection in chickens.</p> </div

Real-time PCR quantification of transcripts coding for constant parts of immunoglobulins in leukocyte subpopulations.

<p>Leukocyte subpopulations were sorted from the spleen of non-infected (N), infected (I) and vaccinated and infected (V) chickens. The infected chickens were i.v. inoculated on day 42 of life and sacrificed 4 days later. IgLC – λ light immunoglobulin chain. Asterisks indicate a significant difference between the expression in sorted leukocyte subpopulations of infected or vaccinated and infected chickens from the same subpopulation of the non-infected chickens (ANOVA, P<0.05).</p

Expression and induction of avidin, serum amyloid A and trappin-6 in macrophages sorted from spleens of non-infected, infected, and vaccinated and infected chickens.

<p>Squares –3 non-infected chickens; circles –3 infected chickens; triangles –3 vaccinated and infected chickens.</p

Expression of genes encoding constant parts of immunoglobulins in the cecum of 4- and 46-day-old chickens.

<p>NI4 – expression in the cecum of 4-day-old, non-infected chickens, INF4 – expression in the cecum of 4-day-old chickens orally infected with <i>S</i>. Enteritidis on day 1 of life, NI46– expression in the cecum of 46-day-old, non-infected chickens, IFN46– expression in the cecum of 46-day-old chickens orally infected with <i>S</i>. Enteritidis on day 42 of life. Blue columns – IgA transcript, green – Ig λ light chain transcript, yellow – IgM transcript, red – IgY transcript.</p

Influence of avidin and biotin on the ability of <i>S</i>. Enteritidis to adhere to and invade the HD11 chicken macrophage-like cell line.

<p>Parentheses indicate whether the pretreatment with avidin and/or biotin was done on the HD11 cell line or <i>S</i>. Enteritidis. Grey scaling is used to simplify individual group differentiation. When the group of all avidin treated HD11 cells was compared with all the remaining experiments in which the cell culture was not pretreated, the comparison came out as significantly different by a t-test with p<0.05, both in adhesion and invasion assays. Avi – avidin, Bio – biotin, SE – <i>S</i>. Enteritidis.</p