Representing true plant genomes: haplotype-resolved hybrid pepper genome with trio-binning

As sequencing costs decrease and availability of high fidelity long-read sequencing increases, generating experiment specific de novo genome assemblies becomes feasible. In many crop species, obtaining the genome of a hybrid or heterozygous individual is necessary for systems that do not tolerate inbreeding or for investigating important biological questions, such as hybrid vigor. However, most genome assembly methods that have been used in plants result in a merged single sequence representation that is not a true biologically accurate representation of either haplotype within a diploid individual. The resulting genome assembly is often fragmented and exhibits a mosaic of the two haplotypes, referred to as haplotype-switching. Important haplotype level information, such as causal mutations and structural variation is therefore lost causing difficulties in interpreting downstream analyses. To overcome this challenge, we have applied a method developed for animal genome assembly called trio-binning to an intra-specific hybrid of chili pepper (Capsicum annuum L. cv. HDA149 x Capsicum annuum L. cv. HDA330). We tested all currently available softwares for performing trio-binning, combined with multiple scaffolding technologies including Bionano to determine the optimal method of producing the best haplotype-resolved assembly. Ultimately, we produced highly contiguous biologically true haplotype-resolved genome assemblies for each parent, with scaffold N50s of 266.0 Mb and 281.3 Mb, with 99.6% and 99.8% positioned into chromosomes respectively. The assemblies captured 3.10 Gb and 3.12 Gb of the estimated 3.5 Gb chili pepper genome size. These assemblies represent the complete genome structure of the intraspecific hybrid, as well as the two parental genomes, and show measurable improvements over the currently available reference genomes. Our manuscript provides a valuable guide on how to apply trio-binning to other plant genomes

Resistance and susceptibility QTL identified in a rice MAGIC population by screening with a minor‐effect virulence factor from Xanthomonas oryzae

Summary Effective and durable disease resistance for bacterial blight (BB) of rice is a continuous challenge due to the evolution and adaptation of the pathogen, Xanthomonas oryzae pv. oryzae (Xoo), on cultivated rice varieties. Fundamental to this pathogens’ virulence is transcription activator-like (TAL) effectors that activate transcription of host genes and contribute differently to pathogen virulence, fitness or both. Host plant resistance is predicted to be more durable if directed at strategic virulence factors that impact both pathogen virulence and fitness. We characterized Tal7b, a minor-effect virulence factor that contributes incrementally to pathogen virulence in rice, is a fitness factor to the pathogen and is widely present in geographically diverse strains of Xoo. To identify sources of resistance to this conserved effector, we used a highly virulent strain carrying a plasmid borne copy of Tal7b to screen an indica multi-parent advanced generation inter-cross (MAGIC) population. Of 18 QTL revealed by genome-wide association studies and interval mapping analysis, six were specific to Tal7b (qBB-tal7b). Overall, 150 predicted Tal7b gene targets overlapped with qBB-tal7b QTL. Of these, 21 showed polymorphisms in the predicted effector binding element (EBE) site and 23 lost the EBE sequence altogether. Inoculation and bioinformatics studies suggest that the Tal7b target in one of the Tal7b-specific QTL, qBB-tal7b-8, is a disease susceptibility gene and that the resistance mechanism for this locus may be through loss of susceptibility. Our work demonstrates that minor-effect virulence factors significantly contribute to disease and provide a potential new approach to identify effective disease resistance

Broad-spectrum resistance and susceptibility to bacterial blight and bacterial leaf streak of rice

Quantitative trait loci (QTL) that confer broad-spectrum resistance (BSR) have been elusive targets of crop breeding programs. Bacterial leaf streak (BLS) and bacterial blight (BB), caused by Xanthomonas oryzae pv. oryzicola (Xoc) and Xanthomonas oryzae pv. oryzae (Xoo), respectively, are responsible for major losses in rice production in Asia and Africa. Controlling these two diseases is particularly important in Sub-Saharan Africa, where no sources of BSR are available in currently deployed varieties. Our goal is to identify novel, broad-spectrum resistance sources to control BLS and BB in rice, using a Multi-parent Advanced Generation Inter-Cross (MAGIC) population, derived from eight elite indica cultivars. MAGIC populations have an increased level of recombination and provide higher precision and resolution to detect QTL. The MAGIC parents and lines were genotyped and phenotyped in both greenhouse and field conditions by screening with diverse strains of Xoc and Xoo. Using genome-wide association and interval mapping analysis, we identified 37 strain-specific QTL, and 14 QTL effective against multiple X. oryzae strains. From these, three QTL are pathovar-specific and 11 confer resistance to both pathovars. By detecting phenotypic effects of causal alleles, we have identified resources that will facilitate a better understanding of how the involved genes contribute to resistance or susceptibility. Because the MAGIC founders are elite varieties, the BSR QTL identified can be rapidly incorporated into breeding programs to achieve more durable resistance to BLS and BB

Quantitative resistance to bacterial pathogens of rice

Disease resistance is the foundation for managing many plant diseases, because resistant varieties have the strongest impact with minimal environmental effects or cost. Consequently, sources of broad-spectrum resistance (BSR), or resistance that is effective against multiple and/or diverse pathogens is of particular interest. However, achieving BSR depends on having effective resistance sources to introgress into elite germplasm. Multi-parent Advanced Generation Inter-Cross (MAGIC) populations are powerful tools for identifying resistance because they have high levels of recombination and enhanced resolution relative to biparental populations. We screened an indica rice MAGIC population developed from eight elite founders for BSR to diverse strains of the rice bacterial blight and leaf streak pathogens Xanthomonas oryzae pv. oryzae (Xoo) and X. o. pv. oryzicola (Xoc), respectively. In addition, building on our hypothesis that durable disease resistance is attainable by targeting key microbial virulence factors, we screened for resistance to Xoo strains isogenic for the known and common virulence factor TAL7b. A combination of genome-wide association studies and interval mapping analyses revealed a number of loci that conferred BSR to both Xoo and Xoc, as well as resistance targeted at TAL7b. These BSR QTL are excellent sources for durable, broadly effective resistance in the field