Combining BAC and PacBio sequencing to resolve cassava mosaic disease resistance locus CMD2

Cassava provides staple food to an estimated 800 million people worldwide and is the most important source of food in developing countries after maize, rice, and wheat. However, cassava production is severely affected by virus-borne cassava mosaic disease (CMD) in Africa and the Indian subcontinent. Annual yield losses due to CMD are estimated to be 55 million tones. A milestone in molecular breeding to for CMD was the identification of single dominant disease resistance locus CMD2. But recent studies have shown that CMD2 cultivars lost resistance during early stages of somatic embryogenesis in tissue culture. It is therefore necessary to understand the mechanism of CMD2-mediated resistance for its efficient utilization in the cassava improvement. My research aims at combining BAC sequencing, whole genome PacBio sequencing and mapping population of CMD resistant (TME3) and susceptible (60444) accessions to fully sequence, assemble and compare the CMD2 locus. We have generated BAC libraries for both cassava genotypes and screened these libraries to identify CMD2-associatted BACs. We have used the information available from classical marker-assisted breeding studies to develop high quality probes for BAC hybridization. Using this pipeline, we have selected and sequenced 33 BACs on PacBio platform, that cover ~3MB of estimated 8MB CMD2 locus. Further, we have used the latest whole genome assemblies of TME3 and 60444 (accessible from ETH Zurich), to perform comparative analysis. In summary, our results have shown that BAC sequencing is an efficient approach for resolving CMD2, specially in the highly repetitive regions. These results have important implications in understanding CMD and developing a long term strategy to control CMD in farmer fields.