Award details

Use of a self-compatible diploid potato for mutagenesis and forward genetic studies.

Reference	BB/K019090/1
Principal Investigator / Supervisor	Dr Matthew Clark
Co-Investigators / Co-Supervisors
Institution	Earlham Institute
Department	Research Faculty
Funding type	Research
Value (£)	140,317
Status	Completed
Type	Research Grant
Start date	26/03/2014
End date	25/03/2017
Duration	36 months

Abstract

Potato, one of the world's major crop plants, has not been subjected to the types of classical genetic analysis that have been employed in other plants. Being a polyploid outbreeder it has been impossible to generate mutational variation, that in other systems, has played such a critical role in elucidating the mechanisms and processes, which underlie plant developmental and related traits. Genetical studies in potato have been limited to the study of relatively few qualitative traits as well as a large number of continuously varying traits. In this project we will address this issue by using a self-compatible Mexican diploid tuber-bearing species of potato Solanum verrucosum to generate a population of EMS mutagenized plants, with a particular focus on identification of mutations that affect plant development, architecture and tuberization. Pilot experiments conducted at JHI have demonstrated the feasibility of this goal, and a few mutants with drastically altered plant development have been obtained and mapping populations for these are under construction. These mutants will be subjected to further genetic and sequence analysis within this project. To this end we will use a novel method, employing next generation sequencing of mutant and wild type bulks to localize a small number of mutants (up to 5) to a small region of the genome. We will also generate draft genome sequence from the progenitor S. verrucosum genotype (VER54-GW1), which will be mapped to the published DM potato genome assembly, as well as assembled de novo. This will provide a template for mutant mapping and isolation, and in addition, it will provide a valuable set of data for comparative genome analysis in potato, and which will contribute to the 'SOL100' initiative aimed at sequencing 100 Solanaceous plant genomes. One of the existing mutants that has a dwarf phenotype will be tested for its fertility to see whether it is feasible to use this in future mutagenesis studies.

Summary

Studies of the genetics of many plants have benefited greatly from the use of mutants, either spontaneous or artificially induced, in which the function of one or more genes has been lost or modified. Mutagenesis can be achieved by use of mutagenic chemicals or other means (e.g. ionizing radiation), and of course all naturally occurring genetic is due to mutation occurring over long time periods. Mendel's famous experiments with peas utilized naturally occurring mutants differing from the 'wild type' in highly visual, single gene traits. Many crop varieties are themselves mutants. The barley variety Golden Promise is a mutant of an older variety Maythorpe. Mutations can be genetically mapped and various approaches can be used to identify the mutated gene, providing useful information, which can be applied to further research and to plant breeding. The crop plant potato is naturally an outbreeding tetraploid, which makes it extremely difficult to generate the sort of mutational variation that has been so useful in other crop plants. This is because potato has four copies of each gene, and to generate a plant that has a mutated version of all four copies of any one gene is virtually impossible. The same is true to a lesser extent in diploid potatoes which have only two copies of each gene (diploids). For this and other reasons genetic studies in potato have lagged behind other plants, and genetic analysis has had to rely on naturally occurring variation, most of which is manifest as continuous or 'non-discrete' variation. In this project we plan to address this by utilizing a species of potato (Solanum verrucosum) that is both diploid and which is a natural inbreeder, allowing the construction of 'homozygous' genotypes, where both copies of each gene are identical. In pilot experiments we have shown that, using S. verrucosum, it is feasible to generate populations of mutant plants that can greatly benefit genetic studies, which will have downstream impact on potato improvement. The mutant panel will be assessed for variation in traits relevant to potato breeding: tuber characteristics, plant architecture traits, tuber sprouting etc and a panel of ~100 interesting mutants will be selected for further study. In pilot experiments a few interesting mutants have been identified and these will be studied within this project. Some of these mutants including one having a clear 'dwarfing' phenotype will used to test a novel approach for isolating the gene that has been mutated. This will also entail generating a draft genome sequence of the genotype used for the mutagenesis. The dwarf mutant will also be tested for its fertility to see whether it is feasible to use this in future mutagenesis studies. The outcomes of this project will be (1) The establishment of the first ever mutant collection of potato. (2) The elucidation of a draft genome sequence of the 'base genotype' of Solanum verrucosum. (3) The identification of candidate genes and mutated alleles for one or more of the mutants identified.

Impact Summary

Who will benefit from this research? The proposal is directly relevant to the BBSRC priority area in Crop Science/Food Security and Living with Environmental Change. The project also builds on BBSRC investment in sequencing the potato genome by generating a draft sequence of a strategically important diploid wild species with unusual biological properties, and to generate a scientific first for potato - a panel of genetic mutants that can be mapped and used to dissect potato biology. Potato researchers will thus benefit directly. In the longer term, potato breeders and eventually farmers stand to benefit. The staff employed on the project will benefit by gaining experience in a completely new set of activities. While TGAC staff have extensive experience of carrying out genome assemblies this will be their first potato genome sequencing project, and it will be interesting to see how well the large families of closely related genes and pseudogenes e.g NB-LRR genes assemble. Again while sequence variation analysis is routine, and TGAC have used the SHOREMAP software before, this was on test Arabidopsis data, real life data from a larger genome species will be an interesting test of the technique. How will they benefit from this research? TGAC staff will benefit from using new informatics tools and tackling new biological problems. They will have to meet project deadlines, and become more connected with the potato researchers and learn more about the underlying potato biology and genetics. Potato researchers will benefit by having access to the new genome sequence, plus the mutant resources generated and the methods used to produce and clone them. The increased focus towards 'monogenic trait biology' will also be a benefit to potato scientists who have customarily worked on the analysis of quantitative traits, which has allowed few opportunities for experimental science. For such scientists it is often quite frustrating to find that variation in a complex trait is controlled by several loci of small effect, and analysis ends at the QTL mapping stage! Having the opportunity to work with discontinuous trait variation would open up new scientific vistas that would enable real experimental science to be performed. Potato researchers, breeders and eventually farmers will be aided by an improved understanding of the potato biology derived from this and future projects. We believe that this project forms an excellent pilot for future potato species and variety resequencing projects which will allow a better understanding of potato biology: plant shape, tuberisation, yield, ability to thrive in a variety of environments and survive pathogens e.g. blight. The project will provide multi-disciplinary training activities for all staff employed on the project, delivering scientists with a high level of scientific and communication skills. The nature of the work will entail the TGAC researchers acquiring new skills such as using the latest genome assembly tools to combine short and long read technologies, and using bulk segregant methods to clone forward genetic mutants. Because we place great importance on excellent communication between teams, and true understanding of how best to meld genetics and genomics together to solve biological problems, TGAC will host the JHI PDRA for two months, training them in informatics skills to use the genome sequence and identify candidate SNPs from the SHOREMAP technique. This exchange of knowledge and skills is bi-directional, the PDRA will share their deep knowledge of the existing potato genome, genetics and biology to highlight the best genome assembly and how to make it more useful for potato researchers. For example the PDRA will use the genome browser and other tools to highlight areas of the genome that are difficult to assemble or particularly important e.g. the NB-LRR genes and also genome browser tracks that can be used to better highlight features for biologists.

Committee	Research Committee B (Plants, microbes, food & sustainability)
Research Topics	Crop Science, Plant Science
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

Associated awards:

BB/K019325/1 Use of a self-compatible diploid potato for mutagenesis and forward genetic studies.

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