RESUMO
BACKGROUND: In this study, whole genome re-sequencing of rust resistant soybean genotype EC241780 was performed to understand the genomic landscape involved in the resistance mechanism. METHODS: A total of 374 million raw reads were obtained with paired-end sequencing performed with Illumina HiSeq 2500 instrument, out of which 287.3 million high quality reads were mapped to Williams 82 reference genome. Comparative sequence analysis of EC241780 with rust susceptible cultivars Williams 82 and JS 335 was performed to identify sequence variation and to prioritise the candidate genes. RESULTS: Comparative analysis indicates that genotype EC241780 has high sequence similarity with rust resistant genotype PI 200492 and the resistance in EC241780 is conferred by the Rpp1 locus. Based on the sequence variations and functional annotations, three genes Glyma18G51715, Glyma18G51741 and Glyma18G51765 encoding for NBS-LRR family protein were identified as the most prominent candidate for Rpp1 locus. CONCLUSION: The study provides insights of genome-wide sequence variation more particularly at Rpp1 loci which will help to develop rust resistant soybean cultivars through efficient exploration of the genomic resource.
RESUMO
Mungbean yellow mosaic India virus (MYMIV) belonging to the family Geminiviridae and the genus Begomovirus is a severe pathogen of tropical legumes including soybean. The absence of genetically mapped loci conferring resistance together with the genetic diversity of begomoviruses infecting soybean warrants the utilization of RNA interference (RNAi) technology to develop virus resistance. However, viral suppressors of RNAi (VSRs) reduce the effectiveness of RNA silencing. Here, we report the effectiveness of Agrobacterium-mediated transient expression of shRNA, targeting a conserved region of AC2 ORF (a VSR) of MYMIV, in conferring virus resistance in soybean. Transient expression of shRNA showed progressive reduction of the viral titre estimated by the MYMIV-derived AC2 gene copy numbers from the initial inoculum by approximately 80-fold 20 days post-application. In addition, the newly emerging leaves exhibited symptom recovery. Thus, this study proves that AC2 of MYMIV is a potent target gene for obtaining RNAi-mediated virus resistance in soybean. Agro-infiltration-based delivery of shRNA was an efficient means of gene silencing and could pave way for the development of transgenic virus-resistant soybean genotype.
RESUMO
Mungbean yellow mosaic India virus (MYMIV) infecting soybean and other legumes causes yellow mosaic disease (YMD). Evaluation of soybean genotypes for YMD resistance involves field screening at disease hot spots or in a protected environment using infectious clones or viruliferous whiteflies as sources of virus inocula. Development of efficient virus inoculation and quantification protocols to screen soybean genetic stocks against YMD is imperative for breeding resistant varieties. Binary plasmids harbouring complete, tandem dimeric genomic components DNA A and DNA B of MYMIV-soybean isolate were engineered. The infectivity of the clones was demonstrated in soybean genotypes JS335 and UPSM534 that display contrasting YMD resistance. As a follow-up, soybean germplasm lines, breeding lines, and representative cultivars that were initially screened at an YMD hot-spot were then subjected to Agrobacterium-based infection with MYMIV. Quantitative real time polymerase chain reaction (qRT-PCR) based copy number analysis of MYMIV genomic components allowed soybean genotypes to be classified into three discrete categories; resistant, moderately resistant and susceptible to the viral infection. Thus, a soybean germplasm disease screening system based on agro-infection and qRT-PCR based quantification of MYMIV was developed to facilitate breeding YMD resistant soybean. The implications of this study for obtaining YMD resistant soybean cultivars are discussed.