Integrating CRISPR-Cas and Next Generation Sequencing in Plant Virology.

Plant pathology has been revolutionized by the emergence and intervention of next-generation sequencing technologies (NGS) which provide a fast, cost-effective, and reliable diagnostic for any class of pathogens. NGS has made tremendous advancements in the area of research and diagnostics of plant infecting viromes and has bridged plant virology with other advanced research fields like genome editing technologies. NGS in a broader perspective holds the potential for plant health improvement by diagnosing and mitigating the new or unusual symptoms caused by novel/unidentified viruses. CRISPR-based genome editing technologies can enable rapid engineering of efficient viral/viroid resistance by directly targeting specific nucleotide sites of plant viruses and viroids. Critical genes such as eIf (iso) 4E or eIF4E have been targeted via the CRISPR platform to produce plants resistant to single-stranded RNA (ssRNA) viruses. CRISPR/Cas-based multi-target DNA or RNA tests can be used for rapid and accurate diagnostic assays for plant viruses and viroids. Integrating NGS with CRISPR-based genome editing technologies may lead to a paradigm shift in combating deadly disease-causing plant viruses/viroids at the genomic level. Furthermore, the newly discovered CRISPR/Cas13 system has unprecedented potential in plant viroid diagnostics and interference. In this review, we have highlighted the application and importance of sequencing technologies on covering the viral genomes for precise modulations. This review also provides a snapshot vision of emerging developments in NGS technologies for the characterization of plant viruses and their potential utilities, advantages, and limitations in plant viral diagnostics. Furthermore, some of the notable advances like novel virus-inducible CRISPR/Cas9 system that confers virus resistance with no off-target effects have been discussed.

Tweaking genome-editing approaches for virus interference in crop plants.

Plant viruses infect various economically important crops and cause a serious threat to agriculture. As of now, conventional strategies employed are inadequate to circumvent the proliferation of rapidly evolving plant viruses. In this regard, recent advancement in genome-editing approach looks promising to produce plants resistant to DNA/RNA virus infections. Clustered regularly interspaced palindromic repeats (CRISPR) system has been emerged as a promising genome-editing tool that has received special interest because of its ease, competence and reproducibility. Recent studies have demonstrated that CRISPR/Cas9 system has great potential to confer plant immunity by either directly targeting or cleaving the viral genome in both RNA and DNA viruses. Similarly, the approach can be used for targeting the host susceptibility genes more particularly in case of RNA viruses. In the present review, different approaches and strategies being used to improve plant resistance against devastating viruses are discussed in view of recent advances in CRISPR systems. This review also describes the major pitfalls of CRISPR/Cas9 system that utilizes highly efficient and novel platforms to engineer interference to single and multiple plant RNA viruses.

Assessment of variability in lignan and fatty acid content in the germplasm of Sesamum indicum L.

Information on the variability available in lignan and fatty acid content in the oilseed crop of Sesamum indicum has been limited. This article presents and discusses the composition, quantity, and variability available for the two traits in the sesame germplasm that are grown in diverse agro climatic regions of India. HPLC and GC analysis of sesame seeds harvested over a period of three crop seasons revealed a considerable amount of variability in lignan and fatty acids. The antioxidant lignans sesamol, sesamin and sesamolin were observed to be in the range of 0.16-3.24, 2.10-5.98 and 1.52-3.76 mg/g of seed, respectively. Similarly oleic and linoleic acids, respectively, have ranged from 34.71 to 45.61% and 38.49 to 49.60%. The black sesame seeds were found rich in sesamin, sesamolin, total lignan content and oleic acid and are thus identified nutritionally and pharmaceutically more important than white and brown seeds. Pearson statistics showed a strong correlation between the components within a particular trait and also some correlation was found between the traits. The study revealed promising cultivars for use in sesame breeding aimed at improving lignan and fatty acid contents, and can be thus directly used in human foods, nutrition, health and welfare.

Genetic diversity and population structure of Cucumis sativus L. by using SSR markers.

The genetic variation, marker attributes and population structure was assessed in 104 genotypes of cucumber using 23 SSR primer pairs. The total number of alleles produced was 67 with an average of 2.91 per locus. Allele frequency was in the range of 0.215 to 0.561 with mean value of 0.403, polymorphic information content ranged from 0.158 to 0.495 with the mean of 0.333, marker index ranged from 0.316 to 1.54 with an average value of 0.954 and resolving power ranged from 0.346 to 2.692 with mean of 1.392. The maximum allele frequency was reported with primer SSR65, whereas the maximum value of polymorphic information content and resolving power was found with SSR61 and the maximum value of marker index was reported with SSR60. Jaccard's similarity coefficient ranged from 0.07 to 0.897 with maximum similarity between genotype G40 and G41 and minimum between G16 and G20, and G16 and G100. Clustering and PCA grouped the genotypes in two clusters, and majority of them were found in cluster B. The population structure analysis also showed two major populations, in which 47 genotypes were found in population 1, 39 genotypes in population 2, whereas remaining 18 genotypes were admixtures. The study provides researchers a valuable information for genotype identification, gene mapping, molecular breeding, and future exploration of cucumber germplasm in India and other major cucumber growing countries.

Comparative assessment of genetic diversity in Sesamum indicum L. using RAPD and SSR markers.

Sesame (Sesamum indicum L.) is an ancient oilseed crop known for its nutty seeds and high-quality edible oil. It is an unexplored crop with a great economic potential. The present study deals with assessment of genetic diversity in the crop. Twenty two RAPD and 18 SSR primers were used for analysis of the 47 different sesame accessions grown in different agroclimatic zones of India. A total of 256 bands were obtained with RAPD primers, of which 191 were polymorphic. SSR primers gave 64 DNA bands, of which all of were polymorphic. The Jaccard's similarity coefficient of RAPD, SSR, and pooled RAPD and SSR data ranged from 0.510 to 0.885, 0.167 to 0.867, and 0.505 to 0.853, respectively. Maximum polymorphic information content was reported with SSRs (0.194) compared to RAPDs (0.186). Higher marker index was observed with RAPDs (1.426) than with SSRs (0.621). Similarly, maximum resolving power was found with RAPD (4.012) primers than with SSRs (0.884). The RAPD primer RPI-B11 and SSR primer S16 were the most informative in terms of describing genetic variability among the varieties under study. At a molecular level, the seed coat colour was distinguishable by the presence and absence of a group of marker amplicon/s. White and brown seeded varieties clustered close to each other, while black seeded varieties remained distanced from the cluster. In the present study, we found higher variability in Sesamum indicum L. using RAPD and SSR markers and these could assist in DNA finger printing, conservation of germplasm, and crop improvement.

Lignans of sesame: purification methods, biological activities and biosynthesis--a review.

Lignans are a group of compounds consisting of dimers of phenyl propane units. They are found in diverse forms distributed in a variety of plants. Sesame lignans in particular are obtained from Sesamum indicum, a highly prized oilseed crop cultivated widely in many countries in the east. The plant is the main source of clinically important antioxidant lignans such as sesamin, sesamolin, sesaminol and sesamol. These lignans exhibit antihypertensive, anticancerous and hypocholesterolemic activities as well especially in humans due to which they have become compounds of tremendous research interest in recent times. Sesamin is synthesized from shikimic acid through phenylpropanoid pathway and metabolised into enterolignans which play a pivotal role in protection against several hormone related diseases. In this paper we present an overview of current status of research on sesame lignans with respect to the analytical methods employed, the biological activities and biosynthesis of sesame lignans.