Speed Breeding Opportunities and Challenges for Crop Improvement

Journal of Plant Growth Regulation - Crop improvement in light of the rapidly changing climate and the increasing human population continues to be one of the primary concerns for researchers across...     

Discriminant haplotypes of avirulence genes of Phytophthora sojae lead to a molecular assay to predict phenotypes

The soybean–Phytophthora sojae interaction operates on a gene-for-gene relationship, where the product of a resistance gene (Rps) in the host recognizes that of an avirulence gene (Avr) in the pathogen to generate an incompatible reaction. To exploit this form of resistance, one must match with precision the appropriate Rps gene with the corresponding Avr gene. Currently, this association is evaluated by phenotyping assays that are labour-intensive and often imprecise. To circumvent this limitation, we sought to develop a molecular assay that would reveal the avirulence allele of the seven main Avr genes (Avr1a, Avr1b, Avr1c, Avr1d, Avr1k, Avr3a, and Avr6) in order to diagnose with precision the pathotypes of P. sojae isolates. For this purpose, we analysed the genomic regions of these Avr genes in 31 recently sequenced isolates with different virulence profiles and identified discriminant mutations between avirulence and virulence alleles. Specific primers were designed to generate amplicons of a distinct size, and polymerase chain reaction conditions were optimized in a final assay of two parallel runs. When tested on the 31 isolates of known virulence, the assay accurately revealed all avirulence alleles. The test was further assessed and compared to a phenotyping assay on 25 isolates of unknown virulence. The two assays matched in 97% (170/175) of the interactions studied. Interestingly, the sole cases of discrepancy were obtained with Avr3a, which suggests a possible imperfect interaction with Rps3a. This molecular assay offers a powerful and reliable tool to exploit and study with greater precision soybean resistance against P. sojae.     

Genome-wide distribution and organization of microsatellites in plants: an insight into marker development in Brachypodium

Plant genomes are complex and contain large amounts of repetitive DNA including microsatellites that are distributed across entire genomes. Whole genome sequences of several monocot and dicot plants that are available in the public domain provide an opportunity to study the origin, distribution and evolution of microsatellites, and also facilitate the development of new molecular markers. In the present investigation, a genome-wide analysis of microsatellite distribution in monocots (Brachypodium, sorghum and rice) and dicots (Arabidopsis, Medicago and Populus) was performed. A total of 797,863 simple sequence repeats (SSRs) were identified in the whole genome sequences of six plant species. Characterization of these SSRs revealed that mono-nucleotide repeats were the most abundant repeats, and that the frequency of repeats decreased with increase in motif length both in monocots and dicots. However, the frequency of SSRs was higher in dicots than in monocots both for nuclear and chloroplast genomes. Interestingly, GC-rich repeats were the dominant repeats only in monocots, with the majority of them being present in the coding region. These coding GC-rich repeats were found to be involved in different biological processes, predominantly binding activities. In addition, a set of 22,879 SSR markers that were validated by e-PCR were developed and mapped on different chromosomes in Brachypodium for the first time, with a frequency of 101 SSR markers per Mb. Experimental validation of 55 markers showed successful amplification of 80% SSR markers in 16 Brachypodium accessions. An online database 'BraMi' (Brachypodium microsatellite markers) of these genome-wide SSR markers was developed and made available in the public domain. The observed differential patterns of SSR marker distribution would be useful for studying microsatellite evolution in a monocot-dicot system. SSR markers developed in this study would be helpful for genomic studies in Brachypodium and related grass species, especially for the map based cloning of the candidate gene(s).     

Identification of major quantitative trait loci qSBR11-1 for sheath blight resistance in rice

Sheath blight caused by Rhizoctonia solani Kühn is one of the important diseases of rice, resulting in heavy yield loss in rice every year. No rice line resistant to sheath blight has been identified till date. However, in some rice lines a high degree of resistance to R. solani has been observed. An indica rice line, Tetep, is a well documented source of durable and broad spectrum resistance to rice blast as well as quantitative resistance to sheath blight. The present study identified genetic loci for quantitative resistance to sheath blight in rice line Tetep. A mapping population consisting of 127 recombinant inbred lines derived from a cross between rice cultivars HP2216 (susceptible) and Tetep (resistant to sheath blight) was evaluated for sheath blight resistance and other agronomic traits for 4 years across three locations. Based on sheath blight phenotypes and genetic map with 126 evenly distributed molecular markers, a quantitative trait loci (QTLs) contributing to sheath blight resistance was identified on long arm of chromosome 11. Two QTL mapping approaches i.e., single marker analysis and composite interval mapping in multi environments were used to identify QTLs for sheath blight resistance and agronomical traits. The QTL qSBR11-1 for sheath blight resistance was identified between the marker interval RM1233 (26.45 Mb) to sbq33 (28.35 Mb) on chromosome 11. This region was further narrowed down to marker interval K39516 to sbq33 (~0.85 Mb) and a total of 154 genes were predicted including 11 tandem repeats of chitinase genes which may be responsible for sheath blight resistance in rice line Tetep. A set of 96 varieties and a F₂ population were used for validation of markers linked to the QTL region. The results indicate that there is very high genetic variation among varieties at this locus, which can serve as a starting point for allele mining of sheath blight resistance.     

Identification of aquaporins and deciphering their role under salinity stress in pomegranate (Punica granatum)

Journal of Plant Biochemistry and Biotechnology - Aquaporins (AQPs), pore-forming proteins, are known to involve in the transport of water and many other small solutes, and play a diverse role in...     

Nitric oxide and hydrogen sulfide crosstalk during heavy metal stress in plants

Gases such as ethylene, hydrogen peroxide (H₂O₂), nitric oxide (NO), carbon monoxide (CO) and hydrogen sulfide (H₂S) have been recognized as vital signaling molecules in plants and animals. Of these gasotransmitters, NO and H₂S have recently gained momentum mainly because of their involvement in numerous cellular processes. It is therefore important to study their various attributes including their biosynthetic and signaling pathways. The present review provides an insight into various routes for the biosynthesis of NO and H₂S as well as their signaling role in plant cells under different conditions, more particularly under heavy metal stress. Their beneficial roles in the plant's protection against abiotic and biotic stresses as well as their adverse effects have been addressed. This review describes how H₂S and NO, being very small-sized molecules, can quickly pass through the cell membranes and trigger a multitude of responses to various factors, notably to various stress conditions such as drought, heat, osmotic, heavy metal and multiple biotic stresses. The versatile interactions between H₂S and NO involved in the different molecular pathways have been discussed. In addition to the signaling role of H₂S and NO, their direct role in posttranslational modifications is also considered. The information provided here will be helpful to better understand the multifaceted roles of H₂S and NO in plants, particularly under stress conditions.     

A precise spacing between the NPA domains of aquaporins is essential for silicon permeability in plants

The controversy surrounding silicon (Si) benefits and essentiality in plants is exacerbated by the differential ability of species to absorb this element. This property is seemingly enhanced in species carrying specific nodulin 26‐like intrinsic proteins (NIPs), a subclass of aquaporins. In this work, our aim was to characterize plant aquaporins to define the features that confer Si permeability. Through comparative analysis of 985 aquaporins in 25 species with differing abilities to absorb Si, we were able to predict 30 Si transporters and discovered that Si absorption is exclusively confined to species that possess NIP‐III aquaporins with a GSGR selectivity filter and a precise distance of 108 amino acids (AA) between the asparagine–proline–alanine (NPA) domains. The latter feature is of particular significance since it had never been reported to be essential for Si selectivity. Functionality assessed in the Xenopus oocyte expression system showed that NIPs with 108 AA spacing exhibited Si permeability, while proteins differing in that distance did not. In subsequent functional studies, a Si transporter from poplar mutated into variants with 109‐ or 107‐AA spacing failed to import, and a tomato NIP gene mutated from 109 to 108 AA exhibited a rare gain of function. These results provide a precise molecular basis to classify higher plants into Si accumulators or excluders.     

An Improved Genotyping by Sequencing (GBS) Approach Offering Increased Versatility and Efficiency of SNP Discovery and Genotyping

Highly parallel SNP genotyping platforms have been developed for some important crop species, but these platforms typically carry a high cost per sample for first-time or small-scale users. In contrast, recently developed genotyping by sequencing (GBS) approaches offer a highly cost effective alternative for simultaneous SNP discovery and genotyping. In the present investigation, we have explored the use of GBS in soybean. In addition to developing a novel analysis pipeline to call SNPs and indels from the resulting sequence reads, we have devised a modified library preparation protocol to alter the degree of complexity reduction. We used a set of eight diverse soybean genotypes to conduct a pilot scale test of the protocol and pipeline. Using ApeKI for GBS library preparation and sequencing on an Illumina GAIIx machine, we obtained 5.5 M reads and these were processed using our pipeline. A total of 10,120 high quality SNPs were obtained and the distribution of these SNPs mirrored closely the distribution of gene-rich regions in the soybean genome. A total of 39.5% of the SNPs were present in genic regions and 52.5% of these were located in the coding sequence. Validation of over 400 genotypes at a set of randomly selected SNPs using Sanger sequencing showed a 98% success rate. We then explored the use of selective primers to achieve a greater complexity reduction during GBS library preparation. The number of SNP calls could be increased by almost 40% and their depth of coverage was more than doubled, thus opening the door to an increase in the throughput and a significant decrease in the per sample cost. The approach to obtain high quality SNPs developed here will be helpful for marker assisted genomics as well as assessment of available genetic resources for effective utilisation in a wide number of species.