RVE2, a new regulatory factor in jasmonic acid pathway, orchestrates resistance to Verticillium wilt.

Verticillium dahliae, one of the most destructive fungal pathogens of several crops, challenges the sustainability of cotton productivity worldwide because very few widely-cultivated Upland cotton varieties are resistant to Verticillium wilt (VW). Here, we report that REVEILLE2 (RVE2), the Myb-like transcription factor, confers the novel function in resistance to VW by regulating the jasmonic acid (JA) pathway in cotton. RVE2 expression was essentially required for the activation of JA-mediated disease-resistance response. RVE2 physically interacted with TPL/TPRs and disturbed JAZ proteins to recruit TPL and TPR1 in NINJA-dependent manner, which regulated JA response by relieving inhibited-MYC2 activity. The MYC2 then bound to RVE2 promoter for the activation of its transcription, forming feedback loop. Interestingly, a unique truncated RVE2 widely existing in D-subgenome (GhRVE2D) of natural Upland cotton represses the ability of the MYC2 to activate GhRVE2A promoter but not GausRVE2 or GbRVE2. The result could partially explain why Gossypium barbadense popularly shows higher resistance than Gossypium hirsutum. Furthermore, disturbing the JA-signalling pathway resulted into the loss of RVE2-mediated disease-resistance in various plants (Arabidopsis, tobacco and cotton). RVE2 overexpression significantly enhanced the resistance to VW. Collectively, we conclude that RVE2, a new regulatory factor, plays a pivotal role in fine-tuning JA-signalling, which would improve our understanding the mechanisms underlying the resistance to VW.

Targeted genome editing in polyploids: lessons from Brassica.

CRISPR-mediated genome editing has emerged as a powerful tool for creating targeted mutations in the genome for various applications, including studying gene functions, engineering resilience against biotic and abiotic stresses, and increasing yield and quality. However, its utilization is limited to model crops for which well-annotated genome sequences are available. Many crops of dietary and economic importance, such as wheat, cotton, rapeseed-mustard, and potato, are polyploids with complex genomes. Therefore, progress in these crops has been hampered due to genome complexity. Excellent work has been conducted on some species of Brassica for its improvement through genome editing. Although excellent work has been conducted on some species of Brassica for genome improvement through editing, work on polyploid crops, including U's triangle species, holds numerous implications for improving other polyploid crops. In this review, we summarize key examples from genome editing work done on Brassica and discuss important considerations for deploying CRISPR-mediated genome editing more efficiently in other polyploid crops for improvement.

Gossypium barbadense and Gossypium hirsutum genomes provide insights into the origin and evolution of allotetraploid cotton.

Allotetraploid cotton is an economically important natural-fiber-producing crop worldwide. After polyploidization, Gossypium hirsutum L. evolved to produce a higher fiber yield and to better survive harsh environments than Gossypium barbadense, which produces superior-quality fibers. The global genetic and molecular bases for these interspecies divergences were unknown. Here we report high-quality de novo-assembled genomes for these two cultivated allotetraploid species with pronounced improvement in repetitive-DNA-enriched centromeric regions. Whole-genome comparative analyses revealed that species-specific alterations in gene expression, structural variations and expanded gene families were responsible for speciation and the evolutionary history of these species. These findings help to elucidate the evolution of cotton genomes and their domestication history. The information generated not only should enable breeders to improve fiber quality and resilience to ever-changing environmental conditions but also can be translated to other crops for better understanding of their domestication history and use in improvement.

Identification of induced mutations in hexaploid wheat genome using exome capture assay.

Wheat is a staple food crop of many countries. Improving resilience to biotic and abiotic stresses remain key breeding targets. Among these, rust diseases are the most detrimental in terms of depressing wheat production. In the present study, chemical mutagenesis was used to induce mutations in the wheat variety NN-Gandum-1. This cultivar is moderately resistant to leaf and yellow rust. The aim of mutagenesis was to improve resistance to the disease as well as to study function of genes conferring resistance to the disease. In the present investigation, a 0.8% EMS dose was found optimum for supporting 45-55% germination of NN-Gandum-1. A total of 3,634 M2 fertile plants were produced from each of the M1 plant. Out of these, 33 (0.91%) and 20 plants (0.55%) showed absolute resistance to leaf and yellow rust, respectively. While 126 (3.46%) and 127 plants (3.49%) exhibited high susceptibility to the leaf and yellow rust, respectively. In the M4 generation, a total of 11 M4 lines (nine absolute resistant and two highly susceptible) and one wild type were selected for NGS-based exome capture assay. A total of 104,779 SNPs were identified that were randomly distributed throughout the wheat sub genomes (A, B and D). Induced mutations in intronic sequences predominated. The highest total number of SNPs detected in this assay were mapped to chr.2B (14,273 SNPs), which contains the highest number of targeted base pairs in the assay. The average mutation density across all regions interrogated was estimated to be one mutation per 20.91 Mb. The highest mutation frequency was found in chr.2D (1/11.7 kb) and the lowest in chr.7D (1/353.4 kb). Out of the detected mutations, 101 SNPs were filtered using analysis criteria aimed to enrich for mutations that may affect gene function. Out of these, one putative SNP detected in Lr21 were selected for further analysis. The SNP identified in chimeric allele (Lr21) of a resistant mutant (N1-252) was located in a NBS domain of chr.1BS at 3.4 Mb position. Through computational analysis, it was demonstrated that this identified SNP causes a substitution of glutamic acid with alanine, resulting in a predicted altered protein structure. This mutation, therefore, is a candidate for contributing to the resistance phenotype in the mutant line. Based on this work, we conclude that the wheat mutant resource developed is useful as a source of novel genetic variation for forward-genetic screens and also as a useful tool for gaining insights into the important biological circuits of different traits of complex genomes like wheat.

QTL mapping for physiology, yield and plant architecture traits in cotton (Gossypium hirsutum L. ) grown under well-watered versus water-stress conditions

Increasing scarcity of irrigation water is a major threat to sustainable production of cotton (Gossypium hirsutum L.). Identifying genomic regions contributing to abiotic stress tolerance will help develop cotton cultivars suitable for water-limited regions through molecular marker-assisted breeding. A molecular mapping F2 population was derived from an intraspecific cross of the drought sensitive G. hirsutum cv. FH-901 and drought tolerant G. hirsutum cv. RH-510. Field data were recorded on physiological traits (osmotic potential and osmotic adjustment); yield and its component traits (seedcotton yield, number of bolls/plant and boll weight); and plant architecture traits (plant height and number of nodes per plant) for F2, F2:3 and F2:4 generations under well-watered versus water-limited growth conditions. The two parents were surveyed for polymorphism using 6500 SSR primer pairs. Joinmap3.0 software was used to construct linkage map with 64 polymorphic markers and it resulted into 35 markers mapped on 12 linkage groups. QTL analysis was performed by composite interval mapping (CIM) using QTL Cartographer2.5 software. In total, 7 QTLs (osmotic potential 2, osmotic adjustment 1, seedcotton yield 1, number of bolls/plant 1, boll weight 1 and plant height 1) were identified. There were three QTLs (qtlOP-2, qtlOA-1, and qtlPH-1) detected only in water-limited conditions. Two QTLs (qtlSC-1 and qtlBW-1) were detected for relative values. Two QTLs (qtlOP-1 and qtlBN-1) were detected for well-watered treatment. Significant QTLs detected in this study can be employed in MAS for molecular breeding programs aiming at developing drought tolerant cotton cultivars.

Detection of single nucleotide polymorphisms in the conserved ESTs regions of Gossypium arboreum

Exploring genetic variation in Gossypium arboreum L. germplasm is useful as it contains many important genes conferring resistance to different stresses. In limited earlier studies, low level of genetic diversity was found by using conventional DNA marker systems which may impede future genome mapping studies. In the present investigation, we explored the extent of Single Nucleotide Polymorphisms (SNP) among 30 conserved regions of Expressed Sequence Tags (EST) of low copy genes between two genotypes of G. arboreum. A total of 27 SNPs including 21 substitutions and 6 Insertions and deletions (Indels) in 7804 bp were found between these genotypes with a frequency of one SNP per 371 bp and one Indel after every 1300 bp. Out of these SNPs, 52 percent were transitions, whilst 48 percent SNPs were transversion. In conclusion, SNPs are expedient markers that can explore polymorphism in highly conserved sequences where other markers are not effective.