Fast, Precise, and Reliable Multiplex Detection of Potato Viruses by Loop-Mediated Isothermal Amplification.

Potato is an important staple food crop in both developed and developing countries. However, potato plants are susceptible to several economically important viruses that reduce yields by up to 50% and affect tuber quality. One of the major threats is corky ringspot, which is a tuber necrosis caused by tobacco rattle virus (TRV). The appearance of corky ringspot symptoms on tubers prior to commercialization results in ≈ 45% of the tubers being downgraded in quality and value, while ≈ 55% are declared unsaleable. To improve current disease management practices, we have developed simple diagnostic methods for the reliable detection of TRV without RNA purification, involving minimalized sample handling (mini), subsequent improved colorimetric loop-mediated isothermal amplification (LAMP), and final verification by lateral-flow dipstick (LFD) analysis. Having optimized the mini-LAMP-LFD approach for the sensitive and specific detection of TRV, we confirmed the reliability and robustness of this approach by the simultaneous detection of TRV and other harmful viruses in duplex LAMP reactions. Therefore, our new approach offers breeders, producers, and farmers an inexpensive and efficient new platform for disease management in potato breeding and cultivation.

Chromosome-scale reference genome assembly of a diploid potato clone derived from an elite variety.

Potato (Solanum tuberosum L.) is one of the most important crops with a worldwide production of 370 million metric tons. The objectives of this study were (1) to create a high-quality consensus sequence across the two haplotypes of a diploid clone derived from a tetraploid elite variety and assess the sequence divergence from the available potato genome assemblies, as well as among the two haplotypes; (2) to evaluate the new assembly's usefulness for various genomic methods; and (3) to assess the performance of phasing in diploid and tetraploid clones, using linked-read sequencing technology. We used PacBio long reads coupled with 10x Genomics reads and proximity ligation scaffolding to create the dAg1_v1.0 reference genome sequence. With a final assembly size of 812 Mb, where 750 Mb are anchored to 12 chromosomes, our assembly is larger than other available potato reference sequences and high proportions of properly paired reads were observed for clones unrelated by pedigree to dAg1. Comparisons of the new dAg1_v1.0 sequence to other potato genome sequences point out the high divergence between the different potato varieties and illustrate the potential of using dAg1_v1.0 sequence in breeding applications.

Precision breeding for novel starch variants in potato.

Potato can be used as a source of modified starches for culinary and industrial processes, but its allelic diversity and tetraploid genome make the identification of novel alleles a challenge, and breeding such alleles into elite lines is a slow and difficult process. An efficient and reliable strategy has been developed for the rapid introduction and identification of new alleles in elite potato breeding lines, based on the ethylmethanesulphonate mutagenesis of dihaploid seeds. Using the granule-bound starch synthase I gene (waxy) as a model, a series of point mutations that potentially affect gene expression or enzyme function was identified. The most promising loss-of-function allele (waxy(E1100)) carried a mutation in the 5'-splice donor site of intron 1 that caused mis-splicing and protein truncation. This was used to establish elite breeding lineages lacking granule-bound starch synthase I protein activity and producing high-amylopectin starch. This is the first report of rapid and efficient mutation analysis in potato, a genetically complex and vegetatively propagated crop.