Plant Breeding Institute

Improving oilseed rape by Cas9-mediated genome editing (OILCAS)

Tahmina2

Background:

Rapeseed is one of the most important oil crops worldwide and achieving high yields has always been the major goal in rapeseed production. The multilocular silique trait has a great potential for the development of high yield varieties of Brassica. Mutation in the CLAVATA signaling pathway genes already showed a promising phenotype in rapeseed. Another major issue in rapeseed cultivation is the pre-harvest seed loss. The fruits, so called siliques, dry out at maturity and become very fragile. Bad weather conditions as strong winds or hail can cause the siliques to break and shed the seeds. This does not only result in yield loss but produces volunteer plants in the next season resulting in quality reduction of the harvest. Knock-out mutations of ALCATRAZ (ALC) produced shatter resistant rapeseed by EMS mutagenesis and the CRISPR-Cas9 system (Braatz et al. 2017, 2018). Another trait of interest is the high amount of glucosinolate content. Glucosinolates are sulfur-containing secondary metabolites, which are common for Brassicas. The effects of these compounds on plant-pest interactions, human health after ingestion, and uptake of rapeseed-based feed by animals are complex. Although breeding has reduced the level of glucosinolates to 8–15 μmol/g in the seeds of modern 00-quality oilseed rape varieties, this amount is still too high for animal feed or human nutrition.

The CRISPR-Cas9 system has recently emerged as an efficient and versatile tool for genome editing in various organisms. However, editing efficiency and use of multiple targets are often limited by the used vector system. A general strategy and platform for precise processing and efficient production of numerous gRNAs using a polycistronic tRNA-processing system would be an advantageous tool in a polyploid crop species like reapeseed.

Objectives:

By knocking out of redundant genes in parallel, we are aiming to produce rapeseed with an increased number of seed chambers, which has a potential to increase seed yield. At the same time, we will target the glucosinolate biosynthesis pathway to reduce antinutritive compounds in rapeseed products. We will apply a vector construct, which allows simultaneous expression of multiple sgRNAs to knock out gene families in rapeseed. Moreover, we expect that the polycistronic tRNA-gRNA molecules are processed by plant innate RNases and yield functional sgRNAs for targeted mutagenesis. We will also test the agronomic performance of two classes of mutants resulting in reduced seed shattering, one obtained by EMS mutagenesis and the other by Cas9-mediated mutagenesis. We expect that CRISPR/Cas modified plants are as fertile and productive as wild type plants while EMS mutants suffer from many background mutations.

Results:

Candidate genes for the multilocular trait will be BnCLV1, BnCLV2 and BnCRN; for the reduction of glucosinolates BnCYP79F and BnMYB28. Crosses to generate a comparable genetic background in BnALC EMS and CRISPR mutants by a novel speed breeding protocol are on the way.

Research Team:

Dr. Tahmina Islam
Dr. Hans-Joachim Harloff
Prof. Dr. Christian Jung

Scientific Partners:

Prof. Dr. rer. nat. Wolfgang Bilger, Ökophysiologie der Pflanzen, University of Kiel

Publications:

Braatz J, Harloff H-J, Jung C (2018) EMS-induced point mutations in ALCATRAZ homoeologs increase silique shatter resistance of oilseed rape (Brassica napus). Euphytica 214:29 doi: 10.1007/s10681-018-2113-7

Braatz J, Harloff H-J, Mascher M, Stein N, Himmelbach A, Jung C (2017) CRISPR-Cas9 targeted mutagenesis leads to simultaneous modification of different homoeologous gene copies in polyploid oilseed rape (Brassica napus). Plant Physiol 174(2):935-942. doi:10.1104/pp.17.00426

Financial Support:

Funding has been provided by the Bundesministerium fur Bildung und Forschung.


Last revision: 29.05.2019                 Responsible for this webpage: Dr. Tahmina Islam