Exploiting seed coat properties to improve uniformity and resilience in Brassica seed vigour

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


A key current goal in plant breeding is to introduce traits that add resilience to climate change. Growers and seed raisers frequently report to the levy board that they have problems with poor or unpredictable germination, even of expensive seed. Seed vigour is determined by the genetics of the crop, by vigour-enhancing formulations applied during seed processing and crucially, by the temperature during seed production. Our evidence suggests that seed traits are the most temperature sensitive in plants, with temperature changes of 1C during seed set capable of making important changes to seed performance. Large seed companies produce seed at specific locations where environment is suited for maximising vigour, but there is still variation from site to site and stochastic temperature fluctuations that affect quality. Increases in weather and climate variation mean that we need to develop the ability to uncouple seed vigour from temperature influences during seed production. This proposal describes a project for breeding new varieties of Brassica with high seed vigour insensitive to the effects of temperature during seed production. Our work has shown that during seed set temperature is sensed by the mother plant and controls progeny seed germination by varying the development and composition of the seed coat. If we alter seed coat development or metabolism genetically, germination remains high regardless of seed set temperature. These processes are widely conserved among all angiosperms, including Brassicas and other vegetable seeds.

Mutagenised populations are important breeding tools in horticulture and have been used historically for novel trait discovery. Here we describe the application of modern state-of-the-art post-genomic technologies to conduct screens of a new mutant B. oleracea population to identify and characterise genes affecting seed coat properties and therefore vigour resilience. We aim to isolate seed coat mutants using simple screens, and using new bioinformatic techniques already available at JIC we can quickly identify causative genes by genome re-sequencing. Using this same population we can also isolate Brassica mutants in specific genes of interest by a process known as TILLING. We know which genes are most important because of extensive work in our lab and others in the closely-related model species Arabidopsis. In this way we can quickly identify Brassica lines in which seed coats and seed vigour resilience are altered compared to laboratory and commercial varieties.

Seed companies also enhance seed vigour by applying chemical formulations in coating to the outside of seeds during processing. These formulations can also include pesticides and fungicides. However, seeds are highly discriminating and uptake of applied chemicals can be as low as 5%, leading to widespread contamination of the environment. If the industry is to continue to benefit from the effects of commercial seed formulations is it clear that uptake into seeds must be improved to minimise the environment impacts of the technology. However, currently there are no techniques available for quantitatively monitoring chemical uptake efficiency into seeds without adding fluorescent labels that also change their properties. A side effect of our seed coat engineering approach is that seed coats control both vigour and permeability to chemicals. Therefore the second part of this proposal is to development an exciting new microscopic technique which can follow the uptake of unlabelled chemicals quantitatively, spatially and in real time into plant tissues, and which also can be used to look inside intact whole seeds (Figure 3). In this way we will be able to see what types of chemical are taken up efficiently into seeds and how this can be altered using genetics. The resulting approach can be used to prioritise development of new chemicals and to show whether more seed permeability can be exploited to maximise chemical uptake.

Technical Summary

Seed vigour is a multi-component trait that is critical for crop performance. Control of dormancy and germination rate are important components of vigour and are dependent on the interaction between plant genetics, the environment during seed production and seed applied agrochemicals. Seeds are highly sensitive to very small temperature changes during seed set, and these are sensed by the mother plant and used to control seed coat traits that affect seed dormancy and seed permeability. Climate change is increasing the unpredictability of vigour in commercial seed lots: therefore the next generation of crops will need high vigour that is uncoupled from effects of temperature variation during seed set. Our work shows that Arabidopsis mutants defective in seed coat development, tannin deposition and suberin deposition have exactly this phenotype. A new B. oleracea TILLING population and new genomic technologies will be exploited in a forward and reverse screening strategy to understand processes controlling seed coat properties in Brassicas, and to generate novel germplasm for breeding programs. Mutants identified from forward screens will be cloned by re-sequencing of bulk segregants and candidate genes confirmed using CRISPR or RNAi, and orthologues of genes known to affect seed permeability in Arabidopsis will be systematically knockout out in B. oleracea. These new alleles will be tested for vigour robustness in laboratory and field trials alongside commercial varieties at sites used for seed production for the UK market. A second strand to this work is the exploitation of seed permeability to enhance chemical uptake into seeds from commercial seed coatings. We will deploy SRS microscopy to analyse spatially, temporally and quantitatively how chemicals with different properties enter seeds with different permeability profiles, and to understand if seed coat permeability engineering has potential to improve chemical uptake into seeds.

Planned Impact

Impacts for the environment
Studies on seed uptake of agrochemicals are few because of limitations of current technology described in this proposal. However, for many widely used compounds uptake is very poor and companies have responded to this simply by increasing dose. The result is that as little as 5% of the applied chemical actually enters the plant, with large quantities accumulating in agro-ecosystems, including soils and watercourses. It has now been shown in recent studies that invertebrate and bird diversity is being affected by seed-applied chemicals. It is becoming clear that the use of many seed-applied compounds may not be sustainable in the long term because these compounds have long half lives in the environment. This project aims to develop seeds that are highly permeable to compounds applied in industrial seed coatings and therefore to markedly improve uptake efficiency. This could lead to lower doses while maintaining effectiveness, and significantly reduce contamination of run-off into soils.
Impacts for seed companies
Seed companies frequently have problems with lot-to-lot variation of seed quality. This occurs because the seeds are produced at multiple sites around the world to ensure continuous supply of fresh seeds, or because stochastic temperature variation at production sites causes quality differences. In some cases this variation can be overcome by post-harvest seed enhancement but this is expensive. Seed companies would benefit now from more predictability in quality, but are concerned that climate change is eroding their ability to reliably produce high quality seeds. The alleles we will isolate and characterise here have the potential to provide robust high quality, even in the face of environmental variation in the seed production conditions. After field trail (objective 3) these can be made available to our industrial collaborator for introgression into elite backgrounds (2-3 years). Because of this link we have a clear route to market and the potential to impact the seed market from 5 years after project end.
Impacts for growers
This project has the potential to impact growers by increasing the reliability of performance of seed they buy from seed companies. Germination of most varieties varies significantly from lot to lot. The new alleles produced in this project have the potential to eliminate much of this variation, improving the predictability and uniformity of seed raising. Increasing uniformity reduces the frequency with which harvest teams visit field sites and increases the proportion of the crop that attains the best grades, adding value for growers.
Training of seed technologists
Seed technologists are individuals trained in the analysing of seed vigour and seed enhancement. Currently there is a global shortage of seed technologists and little training available in the UK. Our program includes extensive experience in both genetics and seed vigour testing and a training secondment in the seed testing lab at Syngenta Vegetable Seeds. These secondments are essential to learn the analytical pipeline, use of software and the rules applied in accredited seed testing labs.
Impact for agrochemical development
Currently there are no good tools for monitoring agrochemical ingress into seeds and thus this is not often considered during development, screening and testing of potential new compounds for commercial seed coatings. SRS microscopy has great potential to be developed into a new tool for the agrochemical industry so that ingress into seeds with different properties can be monitored. In order to facilitate this we will hold a demonstration workshop at the end of the project. If chemicals can be monitored for uptake potential, leads can be screened for uptake potential, and modifications made that would improve uptake into seeds. In future SRS facilities could be built at agrochemical companies.


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Description We have discovered that variation in the environmental tempertaure during seed production causes large variation in seedling growth vigour after germination. We can show that this difference in vigour is related to the tissues surrounding the embryo in the seed, namely the seed coat and a second tissue known as the endosperm. Removing these tissues removes the effect of temperature during seed production on seed vigour. We analysed in detail the response of individual tissues to temperature and identified specific tissues and timepoints at which the seed primarily responds to temperature. We showed that high or low temperatures affects levels of the seed hormone abscisic acid within 1 day of the beginning of the temperature treatment. Finally we identified Brassica mutants in key genes in temperature responses in seeds using the RevGEN TILLING platform at JIC and tested them for use in breeding to speed the breeding of Brassica seeds with higher vigour levels.
Exploitation Route The information can be used in two ways: firstly seed companies can use the information to select seed production sites that give the best quality seeds for their customers. Secondly, when we understand the mechanism we can breed seeds with seed coats with different properties such that their vigour is no longer affected by the temperature during seed production. This increases the reliability of seed quality, which can vary remarkably from lot to lot. Finally the mutants that we discovered could be used in breeding to produce new varieties whose seeds more more resilient to variation in weather during seed production.
Sectors Agriculture, Food and Drink,Other

Description The findings have been used as the basis of two article for famers and growers in AHDB Grower magazine. The project cotributed to a radio didcussion on BBC radio Norfolk food and farming question time. We have used the findings extensively in a collaboration with a major seed company to help inform decision making on the best environmental locations for reliable brassica seed production. In future, the germplasm from the project could be used in brassica breeding for improvements in seed quality.
First Year Of Impact 2018
Sector Agriculture, Food and Drink
Impact Types Economic

Description Brassica Rapeseed And Vegetable Optimisation
Amount £3,514,227 (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
Description Collaboration with Syngenta Seeds 
Organisation Syngenta International AG
Country Switzerland 
Sector Private 
PI Contribution Our contribution is to provide novel genetic materials for field trials by syngenta seeds
Collaborator Contribution Their contribution is to conduct field trials and assist and train for seed analysis.
Impact no outputs yet to report
Start Year 2011
Description Collaboration with the Earlham Institute 
Organisation Earlham Institute
Country United Kingdom 
Sector Academic/University 
PI Contribution We provided expertise in seed germination, and datasets of images of seed germination in a number of species.
Collaborator Contribution Our collaborators used their expertise in machine learning and computer vision to develop algorithms for scoring seed germination
Impact This is an interdisciplinary collaboration, the disciplanes being plant science and artifical intelligence. The outcome is a new automated software tool for scoring seed germination from image stacks.
Start Year 2016
Description collaboration with AHDB 
Organisation Agricultural and Horticulture Development Board
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution We performed research as agreed with the collaborator on brassica seed vigour
Collaborator Contribution The partner provided advice from growers and a platform for dissemination to UK vegetable growers.
Impact no outputs yet to report
Start Year 2015
Description Annual project steering meeting with stakeholders 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Industry/Business
Results and Impact We hosted an annual project meeting where we updated and discussed our work with project stakeholders including syngenta seeds, AHDB, and the Horticulture Innovation partnership. We used industry feedback to modify our priorities.
Year(s) Of Engagement Activity 2017
Description Annual stake holder project meeting 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact We disseminated progress on the project to representatives from syngenta, the horticulture innovation partnership and the agriculture and horticulture development board. They gave advice in return on future project directions.
Year(s) Of Engagement Activity 2017
Description BBC radio norfolk food and farming question time 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Media (as a channel to the public)
Results and Impact I took part in a discussion of innovation in horticulture and agriculture for the local economy, including a discussion of brassica field crops and seed vigour.
Year(s) Of Engagement Activity 2017
Description magazine article in AHDB Grower 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact We prepared an artivcle on the project background for dissmination by AHDB in AHDB Grower. This publication is sent to all UK fruit and vegetable producers.
Year(s) Of Engagement Activity 2018
Description presentation to horticulture and potato initiative meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Industry/Business
Results and Impact gave presentation to assembled group of horticulture industry professionals
Year(s) Of Engagement Activity 2016
Description visit to JIC from Princes foods 
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 We showed our facilities at JIC for seed research and discussed funding of future collaborative projects
Year(s) Of Engagement Activity 2017
Description visit to princes foods 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Industry/Business
Results and Impact Site visit to a large canning company and knowledge exchange. The company are interested in methods to monitor seed coat permeability to improve performance of canning.
Year(s) Of Engagement Activity 2017