BBSRC Renewable Industrial Products from Rapeseed (RIPR) Programme

Lead Research Organisation: University of York
Department Name: Biology

Abstract

Oilseed rape is commercially viable because a co-product can be exploited (a protein-rich feed for animals) in addition to the primary product (oil). However, many other valuable products could be extracted and exploited commercially, adding value to the crop. This opportunity is particularly important for emerging industrial applications of rapeseed oil (such as thermally-stable oil types produced recently by JIC), where the product must compete on price with mineral oil if it is to be successful commercially and hence adopted sufficiently widely to realize the associated environmental benefits. The serial purification of co-products is termed "bio-refining". Emerging opportunities include tocopherols (vitamin E) and phytosterols (cholesterol lowering compounds) from rapeseed oil, waxes (with aphid-repellent properties and those with medical properties) from pod walls and stems, and functional polysaccharides (including high-value stabilisers, surfactants and barriers) from stems. Although the biosynthetic pathways involved in the synthesis of these compounds are known to some extent from studies conducted in species such as Arabidopsis thaliana, the quantitative control of their accumulation (key to yield of the compounds) is not understood. Such products have not been foci for crop improvement by breeders, so there is no improved germplasm and no molecular markers are available to aid breeding. Similarly, the efficiency with which fertilizer is used by rapeseed is poorly understood, but its optimization is essential for the commercial production of substitutes for mineral oil as fertilizer is both expensive and potentially damaging to the environment.

The approach of associating disease or other traits with DNA sequence variation has become a key tool in human genetics and has also been applied successfully in a few plants. The drawback for most crops is that the traditional approach has been expensive. However, recent advances in sequencing technology and genomics has led to the emergence of a technology termed Associative Transcriptomics, which makes this approach accessible for most crops. In this approach, the search for DNA sequence variation is focussed on gene sequences and variation for gene expression is also exploited; both providing very large sets of potential "markers" for trait variation. The technology has been proven recently using a small genetic diversity panel of Brassica napus (the species that includes oilseed rape as one of its crop types) and is ready for scale-up to a full and widely-used genetic diversity panel, i.e. that termed the ASSYST panel. This approach is ideally suited to the initial genetic analysis of traits for which little is know as it quickly enables the estimation of genetic complexity, the development of hypotheses for the control of traits and produces molecular markers that can be used to assist breeding.

The proposed research aims to exploit Associative Transcriptomics to identify genes associated with the control of a range of bio-refining targets and fertilizer use traits. Data will all be made publicly available. The annotations of genes showing either sequence or expression variation associated with trait variation will be examined. Hypotheses will be developed for both the control of the pathways involved and predictive capabilities of molecular markers arising from the identified relationships between gene sequence and/or expression variation and trait variation. These will be tested by the quantitative analysis of traits following the inter-crossing of different plant lines from the collection and/or the selection and testing of plant lines from a population chemically treated to induce gene sequence variation. Using the knowledge gained, mathematical models will be developed to help industry estimate economic and environmental consequences of the development of optimised new cultivars.

Technical Summary

The principal aim of the project is to understand the genetic control, in rapeseed, of the accumulation of compounds representing emerging bio-refining opportunities and fertilizer use efficiency. We will:

1. Establish data models for Brassica transcriptomics and traits. This involves the development of databases and systems enabling the UK Brassica research community to access, analysis and exploit large-scale gene sequence and expression datasets, trait data and marker-trait associations.

2. Determine and make available functional genotypes for Brassica diversity collections. This involves the production of Illumina sequence reads from mRNA extracted from the leaves of each of 600 Brassica accessions (400 B. napus accessions plus 100 accessions of each of B. rapa and B. oleracea), along with the identification and scoring of sequence polymorphisms and quantification of transcript abundance.

3. Improve our understanding of the genetic bases of rapeseed bio-refining traits: tocopherols, phytosterols, waxes and functional polysaccharides, and nutrient use efficiency. This will involve the use of Associative Transcriptomics (a combination of genome-wide association scans, gene expression correlation with trait variation and co-expression network analysis) to identify gene sequence and/or gene expression markers. Hypotheses and markers will be developed for the control of product accumulation. These will be tested by the quantitative analysis of traits following the inter-crossing of plant lines from the collection and/or the selection and testing of plant lines from a "TILLING" population.

4. Develop models for both economic and environmental sustainability of rapeseed. This involves both a cost-benefit analysis for the economic exploitation of co-products from rapeseed and an assessment of the potential environmental impacts, taking into account the potential for improving nutrient use efficiency.

Planned Impact

The proposed research is very well aligned with BBSRC's strategic objectives. Rapeseed is the UK's main vegetable oil crop and of importance for food security (BBSRC's research priority 1), both directly in terms of its production of edible oil and use of residual meal (the material left after extraction of oil from seeds) for feed, and indirectly due to its importance in rotation with cereal crops. Rapeseed is grown on a large scale in the UK (~700,000 Ha in 2011/12) and provides enormous opportunities for bioenergy and industrial biotechnology (BBSRC's research priority 2), particularly for providing renewable substitutes for mineral oil (e.g. modified oil composition for lubricants and biofuels derived from waste straw) and as a source of a range of high value compounds via bio-refining.

Beneficiaries:

The immediate beneficiaries of the research will be the industry partners: Cargill, Biogemma, Limagrain, Monsanto, HGCA, CaseIH and ADAS.

Direct beneficiaries, but on a longer time scale, will be the broader rapeseed breeding and farming industries.

Indirect beneficiaries are the broad range of breeders, farmers and processors around the world working with crops on which the technology will act as an exemplar for rapidly addressing the economic and environmental feasibility of bio-refining.

Benefits:

Cargill will have early access to knowledge of the ranges of quantity and types of co-product that can realistically be accumulated in rapeseed.

Rapeseed breeders will have molecular markers and germplasm to underpin the breeding of rapeseed cultivars with enhanced value from co-products and improved fertilizer use efficiency.

Farmers (represented by HGCA) will be able to sell rapeseed from new cultivars for a higher price per tonne and gain a return from residues such as straw and pod walls, as processors will be able to purify and sell on valuable co-products.

Indirect beneficiaries in other crops will benefit in the same ways as their counterparts in the rapeseed industry.

UK economic performance will benefit as the country's agricultural products will have increased value; similarly for global economic performance.

Public health will benefit as the work will underpin the development of new sources of important compounds with benefits such as dietary antioxidants and lowering LDL cholesterol.

There will be environmental benefits. Directly, alleles that result in improved fertilizer utilization efficiency can be introduced into conventional rapeseed for the production of edible oil. Indirectly, rapeseed lines have recently been developed that produce highly thermo-stable oil that could provide a renewable substitute for mineral oil in a range of applicants such as lubricants and hydraulic fluids. Tocopherols and phytosterols could provide a high value co-product to be extracted from rapeseed oil before such industrial applications, improving the economics of producing such oil.
There are industrial applications for co-products. For example tocopherols are attracting increasing interest as thermal stabilisers in melt polymers (e.g. in the production of LLDPE films) and in lubricants intended for a sensitive environment such as food production. Some of the wax molecular compounds are important semiochemical molecules that help reducing aphid foraging on important food crops. Functional polysaccharides include high value stabilisers, surfactants and barriers.

Publications


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Doheny-Adams T (2017) Development of an efficient glucosinolate extraction method. in Plant methods







 
Description There is extensive variation in the crop species Brassica napus for many co-products of potential use in industrial biotechnology and health. Associative Transcriptomics is effective for identifying molecular markers associated with the genes controlling this variation and in many instances has enabled the identification of candidates for the causative genes themselves.
Exploitation Route They will improve the efficiency of the rapeseed breeding industry. They are likely to underpin the commercial exploitation of a number of co-products from oilseed rape.
Sectors Agriculture, Food and Drink,Chemicals,Energy,Healthcare,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology
 
Description Defra ITT
Amount £556,000 (GBP)
Funding ID CH0104 
Organisation Government of the UK 
Department Department For Environment, Food And Rural Affairs (DEFRA)
Sector Public
Country United Kingdom of Great Britain & Northern Ireland (UK)
Start 01/2015 
End 01/2017
 
Description HEFCE Catalyst
Amount £8,000,000 (GBP)
Organisation Higher Education Funding Council for England (HEFCE) 
Sector Public
Country United Kingdom of Great Britain & Northern Ireland (UK)
Start 05/2015 
End 04/2020
 
Description Manipulating epicuticular wax structure and composition to improve photosynthesis and increase the yield of Brassica oilseed crops
Amount £300,000 (GBP)
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Department Rothamsted Research
Sector Academic/University
Country United Kingdom of Great Britain & Northern Ireland (UK)
Start 02/2015 
End 01/2018
 
Title Brassica Information Portal (BIP) 
Description Manages trait data for UK Brassica research community 
Type Of Material Database/Collection of data 
Year Produced 2017 
Provided To Others? Yes  
Impact Increased awareness of requirements for metadata. 
URL https://bip.earlham.ac.uk/
 
Description 22nd International Symposium on Plant Lipids, Goettingen 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Poster presentation
Development of an ESI-MS/MS method for high throughput profiling of free and esterified sterols and Associative Transcriptomic analysis of seed phytosterol content and composition using a B. napus diversity panel

Richard Broughton1, Lenka Havlickova2, Andrea Harper2, Ian Bancroft2 and Frédéric Beaudoin1
1Biological Chemistry and Crop Protection Department, Rothamsted Research, Harpenden, AL5 2JQ, UK. 2 Department of Biology, University of York, Heslington, YO10 5DD, UK.
Oilseed rape (Brassica napus) is a commercially viable crop owing partly to the seed oil co-products which can be exploited commercially, adding value to the crop. One emerging "bio-refining" opportunity in rapeseed involves the extraction and purification of phytosterols from the oil. Phytosterols are industrially important due to their increasing use as both a functional food component and as nutraceuticals (role in lowering LDL cholesterol). Phytosterols exist in two predominant forms in Brassica napus seeds, free sterols and steryl esters, and it is the latter that is of greater industrial use because of their increased solubility in fat and oil based food products. Although the biosynthetic pathways involved in the synthesis of plant sterols are known to some extent from studies conducted in species such as Arabidopsis thaliana, the quantitative control of their accumulation (key to yield of these compounds) is not understood. In addition, as seed phytosterol content has not been the foci for crop improvement by breeders, there is no improved germplasm or molecular markers available to aid breeding.
We have developed a high throughput ESI-MS/MS method for the compositional and content analysis of seed sterols and steryl esters in a B. napus diversity collection (over 400 accessions representing the ASSYST panel). Here we report the quantitative variations observed and the identification of sequence (SNPs) and gene expression markers (GEMs) associated with these traits in both the A and the C genomes. The strong correlation observed between SNPs and GEMs will greatly facilitate the identification and cloning of the underlying causal genes. Interestingly, we found that very long chain fatty acids are completely excluded from the steryl ester fraction, regardless of the seed fatty acid composition, and that steryl ester content strongly correlates with the seed total sterol content.
Year(s) Of Engagement Activity 2016
URL http://www.eurofedlipid.org/meetings/archive/goettingen2016/
 
Description 7th European Symposium on Plant Lipids (ESPL) in Harpenden 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Presentation of the BBSRC RIPR sLoLa project at the 7th European Symposium on Plant Lipids in Harpenden.
Associative transcriptomic approach to improving the value of co-products in oilseed rape (B. napus)*.
Richard Broughton1, Lenka Havlickova2, Andrea L. Harper2, Ian Bancroft2 and Frédéric Beaudoin1
1 Biological chemistry and crop protection department, Rothamsted research, West Common, Harpenden, AL5 2JQ. 2 Department of Biology, University of York, Heslington, YO10 5DD.

Oilseed rape is commercially viable because a co-product can be exploited (a protein-rich feed for animals) in addition to the primary product (oil). However, many other valuable products could be extracted and exploited commercially, adding value to the crop. This opportunity is particularly important for emerging industrial applications of rapeseed oil where the product must compete on price with mineral oil if it is to be successful commercially and hence adopted sufficiently widely to realize the associated environmental benefits. The serial purification of co-products is termed "bio-refining". Emerging opportunities include tocopherols and phytosterols from rapeseed oil, cuticular waxes from pod walls and stems, and functional polysaccharides from stems. Although the biosynthetic pathways involved in the synthesis of these compounds are known to some extent from studies conducted in species such as Arabidopsis thaliana, the quantitative control of their accumulation (key to yield of the compounds) is not understood. Such products have not been foci for crop improvement by breeders, so there is no improved germplasm and no molecular markers are available to aid breeding. The RIPR* project aims to improve our understanding of the genetic component controlling the quantitative variation observed for selected bio-refining traits of rapeseed (e.g. phytosterol and waxes) building upon knowledge of the biosynthetic pathways developed in species such as A. thaliana. It includes the testing of hypotheses developed using Associative Transcriptomics (an RNAseq-based genome-wide association method focussed on variations in both gene sequence and gene expression), the production of new germplasm and the validation of predictive markers to aid its exploitation by breeders.
We have developed high throughput procedures for quantitative and qualitative analysis of straw/pod wax and seed phytosterol contents in a Brassica diversity collection (circa 600 Brassica accessions: ~400 B. napus accessions representing the ASSYST diversity panel plus B. oleracea, B. rapa and other Brassicaceae to represent broader diversity). Here we report the quantitative variations observed and the identification of both sequence (SNPs) and gene expression markers (GEMs) associated with variations in these traits.
*This work is part of the BBSRC Renewable Industrial Products from Rapeseed (RIPR) Programme (BB/L002124/1 PI: Ian Bancroft)
Year(s) Of Engagement Activity 2015
URL http://www.eurofedlipid.org/meetings/archive/harpenden2015/index.php
 
Description Alcock TD et al. (2015). Identifying genes controlling nutrient uptake and distribution in oilseed rape (Brassica napus). The 14th International Rapeseed Congress, Saskatoon, Canada, July 5-9 2015. [Talk] 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk at scientific conference.
Year(s) Of Engagement Activity 2015
 
Description Alcock TD, Hayden R, Wilson L, Bancroft I, White PJ, Broadley MR, Graham NS (2016) Identifying genes controlling nutrient uptake and distribution in oilseed rape (Brassica napus). Brassica2016 Melbourne, Australia, 3-6 October 2016. [Poster, Talk] 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk at scientific conference.
Year(s) Of Engagement Activity 2016
 
Description Conference (Beijing) 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk on "Genome structural variation in oilseed rape" at 7th International Crop Science Congress, Beijing 15/8/16.
Year(s) Of Engagement Activity 2016
 
Description Fraser A et al. (2015). Improving nitrogen-use efficiency in oilseed rape. The 14th International Rapeseed Congress, Saskatoon, Canada, July 5-9 2015. [Talk] 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk at scientific conference.
Year(s) Of Engagement Activity 2015
 
Description Fraser A et al. (2016) Improving nitrogen use efficiency in oilseed rape (Brassica napus). EMBO Conference - The nitrogen nutrition of plants. Nitrogen 2016, Montpellier, France, 22-26 August 2016. [Talk] 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk at scientific conference.
Year(s) Of Engagement Activity 2016
 
Description Fraser A et al. (2016) Phenotyping to improve nutrient use efficiency in oilseed rape. Canola Innovation Day, Saskatoon, Canada, 1 December 2016. [Talk] 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk at industry/science meeting
Year(s) Of Engagement Activity 2016
 
Description Invited Keynote lecture, 13th Euro Fed Lipid Congress in Florence 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Industry/Business
Results and Impact Oilseed crops as integrated production platforms for food, feed, fuel and renewable industrial feedstock
Frédéric Beaudoin1, Noemi Ruiz-Lopez2, Richard Broughton1, Edgar Cahoon3, Ian Bancroft4 and Johnathan A. Napier1
1 Rothamsted research, Harpenden, AL5 2JQ, UK 2 Instituto de la grasa, Sevilla, Spain 3 Center for Plant Science Innovation, University of Nebraska, Lincoln, USA 4 University of York, Heslington, YO10 5DD, UK.
The world faces considerable challenges including how to produce more biomass for food, feed, fuel and industrial feedstock without significantly impacting on our environment, or increasing our consumption of limited resources such as water or petroleum-derived carbon. This has been described as sustainable intensification. Oleaginous crops have the potential to provide renewable resources for all these commodities, provided they can be engineered to meet end-use requirements and that they can be produced in sufficient quantities to meet current growing world population and industrial demands. This will require more efficient crop production systems but also the development of specialised and high value biotech crops to achieve economic viability.
Although traditional breeding methods have been used successfully to modify the fatty acid composition of oils, metabolic engineering provides a more rapid and direct method for manipulating plant lipid composition. Recent advances in our understanding of the biochemical mechanisms of seed oil biogenesis and the cloning of genes involved in fatty acid and oil metabolic pathways, have allowed the generation of oilseed crops that produce 'designer oils' tailored for specific applications and the conversion of high biomass crops into novel oleaginous crops.
However, improvement of complex quantitative agronomic and seed traits remains more challenging as the underlying genetic determinants are still poorly understood. Technological advances in sequencing and computing have allowed the development of association genetics methods applicable to crops with complex genomes. Associative transcriptomics approaches and high throughput lipidomic profiling can be used to identify the genetic components controlling quantitative variation for lipid-related traits in polyploid crops (like oilseed rape) and provide molecular tools for marker assisted breeding. Examples of traits with potential for bio-refining that can be harvested as co-products in both seeds and vegetative biomass will be presented.
Taken together, progress in plant biotechnology has now made possible the development of multipurpose crops displaying optimised traits for both primary and secondary products. Integrated utilisation of whole oil crops will grow in importance providing renewable resources for health, food and nutrition but also green alternatives for manufacturing and energy production.
Year(s) Of Engagement Activity 2015
URL http://www.eurofedlipid.org/meetings/archive/florence2015/
 
Description Invited Lecture, Oil Crops Research Institute, Chinese Academy of Agricultural Sciences, Wuhan 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Professional Practitioners
Results and Impact Lecture title: Oilseed crops as integrated production platforms for food, feed, fuel and renewable industrial feedstock
Year(s) Of Engagement Activity 2016
 
Description Invited lecture, Institute of Life Sciences Jiangsu University, Zhenjiang, China 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Postgraduate students
Results and Impact Lecture title: Oilseed crops as integrated production platforms for food, feed, fuel and renewable industrial feedstock.
1 h lecture Including a presentation of ther power of Associative transcriptomics for improving the value of co-products in oilseed rape (B. napus).
Year(s) Of Engagement Activity 2016
 
Description Invited lecture, Workshop on Integrative Plant Biology Wuhan, China 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Huazhong (HZAU) - University of Nebraska Lincoln (UNL) Joint workshop on Integrative Plant Biology: Innovations in Crop Improvement to Meet the Global Grand Challenges of the 21st Century. September 2016

Lecture title: Associative transcriptomic approach to improving the value of co-products in oilseed rape (B. napus).
Year(s) Of Engagement Activity 2016
 
Description Invited talk (Abu Dhabi) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk on Associative Transcriptomics for gene discovery presented in Khalifa Centre, Abu Dhabi 19/10/15
Year(s) Of Engagement Activity 2015
 
Description Invited talk (Ludhiana) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk on Associative Transcriptomics and visualization of genome structural evolution in Brassica oilseed crops at Punjab Agricultural University, Ludhiana, India 27/4/16
Year(s) Of Engagement Activity 2016
 
Description Invited talk (New Delhi - NRCPB) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk on Associative Transcriptomics and visualization of genome structural evolution in Brassica oilseed crops at National Research Centre on Plant Biotechnology, New Delhi, India 28/4/16
Year(s) Of Engagement Activity 2016
 
Description Invited talk (New Delhi - UDSC) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk on Associative Transcriptomics and visualization of genome structural evolution in Brassica oilseed crops at University of Delhi South Campus, New Delhi, India 29/4/16
Year(s) Of Engagement Activity 2016
 
Description Invited talk (Wuhan) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk on Visualizing genome structural evolution in Brassica oilseed crops given in Oil Crops Research Institute, Wuhan, China 6/4/16
Year(s) Of Engagement Activity 2016
 
Description White PJ (2016) Bread and potatoes and brassicas - Delivering mineral nutrients for human health. LEAF (Linking Environment and Farming) Technical Event, Balruddery Farm, The James Hutton Institute, 9th June 2016. [Poster, Presentation] 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Talk and poster at technical event
Year(s) Of Engagement Activity 2016