New Approaches for Gene Cloning in Triticeae

The molecular cloning of disease resistance genes traditionally relies on the map-based cloning strategy whereby recombination is used to delimit a gene to a discrete region of the genome. Depending on availability of resources such as genome sequence, genetic maps and mapping populations, this can be a tedious process and potentially futile if the target gene is located in an area of suppressed recombination. An alternative approach that circumvents these problems is mutational genomics. If a gene can be associated with a clear phenotype, then mutating the wildtype material followed by screening for loss of that phenotype provides the basis for genome comparisons. Independent loss-of-resistance mutants most likely are caused by mutations that are distinctly different but located in the target disease resistance gene. The massive genome size of many crop plants is currently prohibitive for whole genome sequencing of multiple mutants. Therefore, we deploy complexity reduction methods such as targeted sequence capture or chromosome flow sorting to minimize the amount of DNA to be sequenced and the volume of data to be analysed. I will provide an update on our pipelines to identify and annotate resistance gene analogues in large-scale omics data, and for cloning candidate disease resistance genes by mutational genomics.