The Brassica napus wall-associated kinase-like (WAKL) gene Rlm9 provides race-specific blackleg resistance.

In plants, race-specific defence against microbial pathogens is facilitated by resistance (R) genes which correspond to specific pathogen avirulence genes. This study reports the cloning of a blackleg R gene from Brassica napus (canola), Rlm9, which encodes a wall-associated kinase-like (WAKL) protein, a newly discovered class of race-specific plant RLK resistance genes. Rlm9 provides race-specific resistance against isolates of Leptosphaeria maculans carrying the corresponding avirulence gene AvrLm5-9, representing only the second WAKL-type R gene described to date. The Rlm9 protein is predicted to be cell membrane-bound and while not conclusive, our work did not indicate direct interaction with AvrLm5-9. Rlm9 forms part of a distinct evolutionary family of RLK proteins in B. napus, and while little is yet known about WAKL function, the Brassica-Leptosphaeria pathosystem may prove to be a model system by which the mechanism of fungal avirulence protein recognition by WAKL-type R genes can be determined.

WheatCRISPR: a web-based guide RNA design tool for CRISPR/Cas9-mediated genome editing in wheat.

BACKGROUND: CRISPR/Cas9 gene editing has become a revolutionary technique for crop improvement as it can facilitate fast and efficient genetic changes without the retention of transgene components in the final plant line. Lack of robust bioinformatics tools to facilitate the design of highly specific functional guide RNAs (gRNAs) and prediction of off-target sites in wheat is currently an obstacle to effective application of CRISPR technology to wheat improvement. DESCRIPTION: We have developed a web-based bioinformatics tool to design specific gRNAs for genome editing and transcriptional regulation of gene expression in wheat. A collaborative study between the Broad Institute and Microsoft Research used large-scale empirical evidence to devise algorithms (Doech et al., 2016, Nature Biotechnology 34, 184-191) for predicting the on-target activity and off-target potential of CRISPR/SpCas9 (Streptococcus pyogenes Cas9). We applied these prediction models to determine on-target specificity and potential off-target activity for individual gRNAs targeting specific loci in the wheat genome. The genome-wide gRNA mappings and the corresponding Doench scores predictive of the on-target and off-target activities were used to create a gRNA database which was used as a data source for the web application termed WheatCRISPR. CONCLUSION: The WheatCRISPR tool allows researchers to browse all possible gRNAs targeting a gene or sequence of interest and select effective gRNAs based on their predicted high on-target and low off-target activity scores, as well as other characteristics such as position within the targeted gene. It is publicly available at https://crispr.bioinfo.nrc.ca/WheatCrispr/ .

A high-contiguity Brassica nigra genome localizes active centromeres and defines the ancestral Brassica genome.

It is only recently, with the advent of long-read sequencing technologies, that we are beginning to uncover previously uncharted regions of complex and inherently recursive plant genomes. To comprehensively study and exploit the genome of the neglected oilseed Brassica nigra, we generated two high-quality nanopore de novo genome assemblies. The N50 contig lengths for the two assemblies were 17.1 Mb (12 contigs), one of the best among 324 sequenced plant genomes, and 0.29 Mb (424 contigs), respectively, reflecting recent improvements in the technology. Comparison with a de novo short-read assembly corroborated genome integrity and quantified sequence-related error rates (0.2%). The contiguity and coverage allowed unprecedented access to low-complexity regions of the genome. Pericentromeric regions and coincidence of hypomethylation enabled localization of active centromeres and identified centromere-associated ALE family retro-elements that appear to have proliferated through relatively recent nested transposition events (<1 Ma). Genomic distances calculated based on synteny relationships were used to define a post-triplication Brassica-specific ancestral genome, and to calculate the extensive rearrangements that define the evolutionary distance separating B. nigra from its diploid relatives.

Genes associated with chloroplasts and hormone-signaling, and transcription factors other than CBFs are associated with differential survival after low temperature treatments of Camelina sativa biotypes.

Winter annual biotypes of Camelina sativa regularly survive after winter conditions experienced in northern regions of the U.S., whereas summer annual biotypes do not. To determine potential molecular mechanisms associated with these biotype differences in survival after low temperature treatments, we examined genetic and transcript variations in both a winter- (Joelle) and a summer- (CO46) biotype. It was determined that as few as one or two dominant genes may control differential survival after low temperature treatments. Of the 1797 genes that were differentially expressed in response to cold in both the winter and summer biotypes many COR genes were identified, indicating that the CBF regulon is functional in both. However, only 153 and 76 genes from Joelle and CO46, respectively, were either differentially expressed or not expressed at all in one biotype versus the other following cold acclimation. We hypothesize that these 229 genes play a significant role in, or are primarily responsive to, differences in survival after freezing between these two biotypes. Promoter analysis provided few clues as to the regulation or these genes; however, genes that were down-regulated specifically in the winter biotype Joelle were enriched with the sequence TGGCCCTCGCTCAC, which is over-represented among genes associated with chloroplasts in Arabidopsis. Additionally, several genes involved in auxin signaling were down-regulated specifically in Joelle. A transcription factor with strong similarity to MYB47, known to be up-regulated by salt, drought, and jasmonic acid, but not cold in Arabidopsis, was essentially off in the freezing sensitive biotype CO46, but was cold-induced in the winter biotype Joelle. Several other transcription factors genes including three with similarity to WRKY70, that may be involved in SA/JA-dependent responses, a HOMEOBOX 6 gene involved in ABA signaling, and two others (NUCLEAR FACTOR Y and CONSTANS-like 2) known to be implicated in photoperiodic flowering were also differentially expressed between the two biotypes.