Exploring knowledge of gene function to combat pod shatter in oilseed rape

Lead Research Organisation: John Innes Centre
Department Name: Crop Genetics

Abstract

Successful domestication of seed crops depends on the plant's ability to hold on to its progeny until harvest. The problem of seed dispersal control in cereals was solved thousands of years ago through simple selective breeding, but remains a serious issue for oilseed rape. The average annual loss experienced by farmers due to premature fruit opening, known as pod shatter is >10%. This loss can exceed >70% under particularly windy conditions, when wet weather delays harvest or if a hailstorm hits the field when the crop is ripe. With an expanding human population and dramatic changes in climate patterns, the challenge to global food production has never been bigger, and to meet the demands it is essential that performance of our major crops be improved. However, the potential yield gain from such efforts cannot be fully realised if the farmer looses a significant part of his crop even before going out to harvest. Fruits from oilseed rape dry out at maturity and open to allow their seeds to be dispersed in a process known as pod shatter. Unfortunately, all the fruits in the field do not dry out at the same time making it difficult for oilseed rape farmers to time their harvest and obtain all the seeds. In addition to a significant yield loss, the prematurely released seeds fall to the ground and germinate to become weeds (volunteers) and contaminate the harvest of the following year. This severely inhibits the crop rotation practice used by many farmers and is therefore also damaging to the environment. Arabidopsis is a small, weedy plant that has been used as a laboratory model system to elucidate a wide range of aspects related to plant growth and development. Despite a dramatic size difference, fruits from Arabidopsis are remarkably similar to fruits from oilseed rape. In the past decade some of the key genetic regulators of fruit opening in Arabidopsis have been identified, and we have shown that these factors also function in species that are closely related to oilseed rape. In the proposed research project, we will exploit our knowledge to control pod shatter directly in high-yielding UK-elite oilseed rape varieties. Specifically we will manipulate and adjust the activity of a particular gene by the isolation of mutant plants and assessment of their performance in pod shatter-resistance tests. We will furthermore use mutated lines to expand our knowledge about the mechanism of fruit opening to enable us to fine-tune the pod shatter trait in future varieties. Particular strengths of this project are our genetic resources and our expertise in all the required technologies to successfully fulfill our objectives. In conclusion, we believe that the presented project will provide substantial benefits for both farmers and consumers, as well as for the environment.

Technical Summary

The aim of this project is to introduce pod shatter resistance into modern oilseed rape varieties by modification of two paralogous genes. We have previously described how the INDEHISCENT (IND) gene regulates shattering in the model plant Arabidopsis and how this knowledge can be transferred to Brassica species such as B. rapa and B. oleracea. Here, resistant lines will be identified via TILLING (Targeting Induced Local Lesions IN Genomes) for mutations in the two B. napus IND genes. Single and double mutants will be characterised for their ability to fine-tune the shattering process and will be backcrossed to elite varieties for immediate use in breeding programmes. In addition to the significant commercial and environmental value of such lines, the experiments that we have planned will expand our knowledge on the molecular mechanism by which IND mediates its function. To this end, we will take advantage of a large allelic series of point mutations in the IND gene in B. rapa (BraA.IND). IND regulates hormone balance at the valve margin, and the mutant lines will be tested for their ability to create an auxin minimum and direct expression of the GA biosynthesis gene GA4. We have previously shown that IND can form homodimers in yeast and heterodimers with a related transcription factor, SPATULA. We will create site-directed mutants in the BraA.IND gene corresponding to those identified in the allelic series to map the domains required for these protein-protein interactions. In this way, the proposed project will drastically reduce and possibly eliminate pod shatter in oilseed rape and will advance our knowledge on fruit tissue-specification while developing tools for further fine-tuning of pod shatter resistance in future varieties.

Planned Impact

Who will benefit from this research and how? Oilseed rape makes a considerable contribution to UK agriculture; currently ~20% to the total crop output per year is devoted to its production (information from The International Agri-Technology Centre). Production is increasing each year with an area of 581,000 ha of oilseed rape sown in 2009 producing 2.0 million tones (compared to 1.5 million tones in 2002). Still the UK imports more than 150,000 tonnes oilseed per year. The yield could be significantly improved by inhibiting premature pod shatter, which leads to annual losses of 15-20% on average but can exceed 70% under adverse weather conditions (see supporting letter from organic farmer, Philip Taylor). Besides resulting in an estimated loss of ~£60 million in the UK per year, the prematurely released seeds fall to the ground and germinate to become weeds (volunteers) and contaminate the harvest of the following year. This severely inhibits the crop rotation practice used by many farmers and is therefore also damaging to the environment. The output of this research will therefore be of value to all industries with interest in crop improvement. The agricultural industry/agronomists: The industry will benefit from the development of technologies to minimise seed loss due to unsynchronised seed dispersal. IP-free pre-breeding material will be made available for the industry with first-hand access to industrial members of CIRC. Farmers: Reduction in seed loss will increase the farmer's yield. The ~10% losses is an estimated average that hides the occasional disasters when strong winds or a hailstorm can cause >70% loss of yield. Importantly, the research proposed here will not only lead to increased yields, but the protection against the effect of adverse weather conditions will also make the yield more predictable. Public: The public would benefit from greater predictability of yields, through greater stability in production costs, which would impact on prices in the shops. There are also obvious environmental benefits using the technology described here. Oilseed rape has emerged as the second largest oilseed crop with an annual worldwide production of 38 million tons of oil and demand is increasing. For this to be sustainable, seed yield needs to be dramatically increased through more efficient breeding programmes while at the same time minimising the amount of fertiliser input in order to protect the environment. I believe the strategy adopted here will contribute significantly towards such a goal. What will be done to ensure that they have the opportunity to benefit from this research? Publications: Results will be published in high-impact scientific journals and the breeding/farming press in a timely fashion. It will also be presented at national and international conferences and trade shows. Collaborations: The PI has strong connections to the breeding industry and Brassica crop improvement programmes. The data and pre-breeding material that we obtain will be of immediate use to these interest groups for example via CIRC dissemination meetings and the Defra-funded Oilseed Rape Genetic Improvement Network (OREGIN) which brings together academic researchers and breeding companies to generate pre-breeding material. Commercialisation: Contacts with industrialists, biotechnologists and related umbrella organisations will be made through CIRC and OREGIN as soon as any exploitable results/materials are generated. The strategy we adopt will allow the IP-free pre-breeding material to be tested both at JIC and by our industry contacts at their sites. We will enquire with the breeding companies about suitable elite lines for backcrossing to facilitate the route to exploitation.
 
Description This project has established a B. napus mutant population as a TILLING platform for reverse genetics.
We have used this resource to successfully produce mutant oilseed rape lines with a reduction in pod shatter. As part of this project, we identified two paralogues of the INDEHISCENT (IND) gene known to be required for fruit dehiscence in Arabidopsis. We identified allelic series of point mutations in both paralogues in B. napus. Whilst the single mutants did not demonstrate any effect on pod shatter compared to wild type, we produced double-mutant combinations some of which exhibited reduced fruit opening in the random impact test. The mutated lines are ready for distribution to our industrial partners within CIRC and are expected to have a significant effect on improving harvestable yield. A manuscript containing an analysis of the phenotypic characteristics of these lines in terms of tissue specification and shatter-potential is under review. This manuscript also presents a new and optimised software package for analysis of random impact test data.
The six-monthly dissemination meetings were excellent opportunities for grant holders to meet members of the industry. Through these meetings we significantly strengthened our interactions with industry. I have no doubt that these interactions were pivotal in being able to design the BRAVO programme with support from industry.
In addition to the potential for crop improvement, this project has contributed to an increase in our understanding of hormonal and genetic regulation of plant organ development. Importantly, it has provided the last step in the most illustrative model-to-crop pipeline example to date. This project is based on fundamental knowledge on Arabidopsis fruit development and the identification of the key components involved. Through proof-of-concept projects in diploid Brassicas, this project demonstrated the power of translation all the way to the crop.
A manuscript reporting the outcome of this grant was published in the journal Plant Reproduction (Stephenson et al., 2019).
Exploitation Route The shatter-resistant oilseed rape lines that we have produced are ready to enter into breeding programmes and the industrial members of the CIRC programme have been notified about their availability. Moreover, this project was pivotal in establishing the best example to date on successful model-to-crop translation. Our work demonstrates the huge potential for such approaches by revealing how knowlege on fruit patterning in Arabidopsis can be translated via proof-of-concept in diploid Brassica species to the B. napus crop species.
Sectors Agriculture, Food and Drink

 
Description Through meetings with the Agriculture and Horticultural Development Board (AHDB), it is clear that there is an increasing interest in including pod shatter assessment of oilseed rape varieties that are on the UK Recommended List. This assessment will be through the use of the Random Impact Test system and the newly developed software associated with the analysis of data, both of which were developed at the JIC. This would be a fantastic outcome in terms of impact and discussions on how best to facilitate this are ongoing between AHDB and JIC. We have enjoyed great interactions with stakeholders in this grant which has helped us to engage more closely with industry. As part of this CIRC grant, we met with stakeholders and other grant holders every 6 months for updates and discussions. In addition, this grant gave me an opportunity to speak about the work on the BBC Radio 4 programme "Farming Today" in Nov 2012. I also gave a presentation at the AgriTech East pollinator event in Oct 2015, which was followed up by an article in the Eastern Daily Press the day after.
First Year Of Impact 2011
Sector Agriculture, Food and Drink,Education
Impact Types Societal

 
Description Presentation of PCGIN, WGIN and ORIGIN impact to DEFRA: provided data to support DEFRA continuing GIN support, engagement with GRU national capability and DEFRA engagement in JIC research.
Geographic Reach National 
Policy Influence Type Gave evidence to a government review
 
Description Research Impact on Policy- visit to review impact of Genetic Improvement Networks
Geographic Reach National 
Policy Influence Type Participation in a advisory committee
 
Description Project with Brandon Biosciences Ltd to test the effect of seaweed extracts on pod shatter in oilseed rape
Amount € 170,000 (EUR)
Funding ID Project ID: 652506 
Organisation European Commission H2020 
Sector Public
Country Belgium
Start 10/2017 
End 09/2018
 
Description Strategic LoLa
Amount £4,392,784 (GBP)
Funding ID BB/P003095/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 01/2017 
End 12/2021
 
Title Random Impact Test version 2.0 
Description The Random Impact Test is to use for assessing pod shatter resistance in oilseed rape. We have further improved the method to also include other members of the Brassicaceae family including Arabidopsis, Capsella and Lepidium. Moreover, newly developed software for curve fitting has allowed more precise and automated calculations. 
Type Of Material Biological samples 
Year Produced 2016 
Provided To Others? Yes  
Impact We have been approached by industry and academics to carry out this assay on a range of material. 
 
Description Testing seaweed extract for combatting pod shatter in oilseed rape 
Organisation Brandon Products Ltd
Country Ireland 
Sector Private 
PI Contribution Testing different formulars of seaweed extract in reducing pod shatter of oilseed rape. Assessing whether the effect occurs via the SHP/IND-mediated pathway by testing the effect of the extracts on gene expression in Arabidopsis fruits.
Collaborator Contribution Production of seaweed extracts and further refinement to identify active compounds.
Impact Seaweed extracts have been tested on several varieties of oilseed rape as well as on reporter lines of Arabidopsis. No effect has been detected on pod shatter resistance, but it does seem in Arabidopsis as if the valve margin auxin response minimum is compromised, suggesting the extract has an effect on the programme leading to seed dispersal.
Start Year 2016
 
Description BRAVO Workshop 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Industry/Business
Results and Impact The BRAVO Workshop aimed to identify future research on oilseed rape and vegetable Brassicas for optimal development and yield. The workshop was effective by allowing JIC scientists to listen to the industry regarding important target traits with the aim of putting together a SLoLa application. The BRAVO SLoLa has now been funded.
Year(s) Of Engagement Activity 2015
 
Description BRAVO stakeholder meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Industry/Business
Results and Impact Annual BRAVO sLoLa stakeholder meetings where members of the breeding/growing industry discuss progress on the BRAVO project.
Year(s) Of Engagement Activity 2018
 
Description Breeders Day 2019 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Industry/Business
Results and Impact This event was designed to allow plant breeders, agronomists and policy makers to find out about some the JIC's latest developments in germplasm for future crops, both wheat and Brassica crops. Early career researchers presented their findings and discussions allowed us to understand key concerns from industry which we can help to address.
Year(s) Of Engagement Activity 2019
 
Description Engagement with Industry- iCASE coordination meeting at Limagrain UK 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Industry/Business
Results and Impact In October 2013 Lars Ostergaard had a meeting with Limagrain UK to coordinate a joint iCASE project on fruit form.
Year(s) Of Engagement Activity 2013
 
Description Engagement with not for profit organisations- visit by Susannah Bolton, AHDB 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Third sector organisations
Results and Impact Visit by Susannah Bolton, AHDB to the JIC to extend the understanding of JIC science that my be relevant to AHDB.
Year(s) Of Engagement Activity 2015
 
Description Engagement with other research organisations and HEIs: Potential Kew and JIC Brassica interactions 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Professional Practitioners
Results and Impact A meeting was organised to discuss a future collaboration between Kew gardens research and JIC brassica molecular agronomy.
Year(s) Of Engagement Activity 2016
 
Description NFU discussion on translation of Brassica research 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Industry/Business
Results and Impact Invited to speak to the South Norfolk NFU group about Brassica research and it's implications for the national farming community. Discussions on our research into pod shatter, flea beetle, temperature effects on oilseed rape and improving yield were well received and sparked many questions and ideas on how best to tackle some of the challenges farmers are facing growing oilseed rape. We also discussed the the technologies e.g. CRISPR we our both using and developing and how this would provide benefits in the years to come. This included a two way exchange of ideas on how best to manage their challenges and what they would find useful in our future research.
Year(s) Of Engagement Activity 2020
 
Description Radio interview- Radio 4 Farming Today 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Interview in the glasshouse among Brassica plants with Anna Hill from BBC Radio 4 Farming Today programme. Specifically describing the BRAVO programme with clear connections and links to the GEN ISP.
Year(s) Of Engagement Activity 2017
 
Description Radio interview- Radio 4 Farming Today 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Media (as a channel to the public)
Results and Impact In October 2012 Lars Ostergaard participated in a radio interview for Radio 4 Farming Today discussing his research findings about pod shatter in oilseed rape.
Year(s) Of Engagement Activity 2012