Genome analysis and avirulence gene cloning using a high-density linkage map for the flax rust fungus

C Anderson1, MA Khan1, CA Jack2, A-M Catanzariti1, A Nemri3, GJ Lawrence3, AR Hardham1, JG Ellis3, PN Dodds3 and DA Jones1

  1. Research School of Biology, The Australian National University, ACT 2601, Australia
  2. ANU Bioinformatics Consultancy, The Australian National University, ACT 2601, Australia
  3. CSIRO Agriculture, Canberra, ACT 2601, Australia

The first genetic linkage map was made over 100 years ago and for the past few decades maps made from molecular markers have been fundamental tools for marker-assisted breeding, quantitative trait loci identification and gene identification by map-based cloning. The advent of next-generation sequencing technologies has revolutionised genetic marker identification, enabling construction of high-density genetic maps containing tens of thousands to millions of sequence-characterised markers. These maps can be used for new purposes, such as anchoring genome assemblies and genome-wide analysis of recombination rate. We have developed a high-density genetic map for the flax rust fungus, Melampsora lini, a plant pathogen and model for understanding the biology of agriculturally important rust fungi. The map comprises 13,412 restriction site-associated DNA sequencing (RADseq) markers in 27 linkage groups that together span 5,860 cM and contain 2,756 recombination bins. The marker sequences were used to anchor 68.9% of the M. lini genome assembly onto the genetic map. The map and anchored assembly were then used to: 1) show that M. lini has a high overall meiotic recombination rate, but recombination distribution is uneven and large coldspots exist; 2) show that substantial genome rearrangements have occurred in spontaneous loss-of-avirulence mutants; and 3) identify the AvrL2 and AvrM14 avirulence genes by map-based cloning.