The phenomenon of RNA interference (RNAi) which involves sequence-specific gene regulation by small non-coding RNAs, i.e., small interfering RNA (siRNA) and microRNA (miRNA) has emerged as one of most powerful approaches for crop improvement. RNAi based on siRNA is one of the widely used tools of reverse genetics which aid in revealing gene functions in many species. This technology has been extensively applied to alter the gene expression in plants with an aim to achieve desirable traits. RNAi has been used for enhancing the crop yield and productivity by manipulating the gene involved in biomass, grain yield and enhanced shelf life of fruits and vegetables. It has also been applied for developing resistance against various biotic (bacteria, fungi, viruses, nematodes, insects) and abiotic stresses (drought, salinity, cold, etc.). Nutritional improvements of crops have also been achieved by enriching the crops with essential amino acids, fatty acids, antioxidants and other nutrients beneficial for human health or by reducing allergens or anti-nutrients. microRNAs are key regulators of important plant processes like growth, development, and response to various stresses. In spite of similarity in size (20–24 nt), miRNA differ from siRNA in precursor structures, pathway of biogenesis, and modes of action. This review also highlights the miRNA based genetic modification technology where various miRNAs/artificial miRNAs and their targets can be utilized for improving several desirable plant traits. microRNA based strategies are much efficient than siRNA-based RNAi strategies due to its specificity and less undesirable off target effects. As per the FDA guidelines, small RNA (sRNA) based transgenics are much safer for consumption than those over-expressing proteins. This review thereby summarizes the emerging advances and achievement in the field of sRNAs and its application for crop improvement.

New advances in crop genetic engineering can significantly pace up the development of genetically improved varieties with enhanced yield, nutrition and tolerance to biotic and abiotic stresses. Genetically modified (GM) crops can act as powerful complement to the crops produced by laborious and time consuming conventional breeding methods to meet the worldwide demand for quality foods. GM crops can help fight malnutrition due to enhanced yield, nutritional quality and increased resistance to various biotic and abiotic stresses. However, several biosafety issues and public concerns are associated with cultivation of GM crops developed by transgenesis, i.e., introduction of genes from distantly related organism. To meet these concerns, researchers have developed alternative concepts of cisgenesis and intragenesis which involve transformation of plants with genetic material derived from the species itself or from closely related species capable of sexual hybridization, respectively. Recombinase technology aimed at site-specific integration of transgene can help to overcome limitations of traditional genetic engineering methods based on random integration of multiple copy of transgene into plant genome leading to gene silencing and unpredictable expression pattern. Besides, recently developed technology of genome editing using engineered nucleases, permit the modification or mutation of genes of interest without involving foreign DNA, and as a result, plants developed with this technology might be considered as non-transgenic genetically altered plants. This would open the doors for the development and commercialization of transgenic plants with superior phenotypes even in countries where GM crops are poorly accepted. This review is an attempt to summarize various past achievements of GM technology in crop improvement, recent progress and new advances in the field to develop improved varieties aimed for better consumer acceptance.

Remarkable diversity in the domain of genome loci architecture, structure of effector complex, array of protein composition, mechanisms of adaptation along with difference in pre-crRNA processing and interference have led to a vast scope of detailed classification in bacterial and archaeal CRISPR/Cas systems, their intrinsic weapon of adaptive immunity. Two classes: Class 1 and Class 2, several types and subtypes have been identified so far. While the evolution of the effector complexes of Class 2 is assigned solely to mobile genetic elements, the origin of Class 1 effector molecules is still in a haze. Majority of the types target DNA except type VI, which have been found to target RNA exclusively. Cas9, the single effector protein, has been the primary focus of CRISPR-mediated genome editing revolution and is an integral part of Class 2 (type II) system. The present review focuses on the different CRISPR types in depth and the application of CRISPR/Cas9 for epigenome modification, targeted base editing and improving traits such as abiotic and biotic stress tolerance, yield and nutritional aspects of tomato breeding.

Oxalate decarboxylase (OXDC) enzyme has immense biotechnological applications due to its ability to decompose anti-nutrient oxalic acid. Flammulina velutipes, an edible wood rotting fungus responds to oxalic acid by induction of OXDC to maintain steady levels of pH and oxalate anions outside the fungal hyphae. Here, we report that upon oxalic acid induction, a calmodulin (CaM) like protein-FvCaMLP, interacts with the OXDC promoter to regulate its expression. Electrophoretic mobility shift assay showed that FvCamlp specifically binds to two non-canonical E-box elements (AACGTG) in the OXDC promoter. Moreover, substitutions of amino acids in the EF hand motifs resulted in loss of DNA binding ability of FvCamlp. F. velutipes mycelia treated with synthetic siRNAs designed against FvCaMLP showed significant reduction in FvCaMLP as well as OXDC transcript pointing towards positive nature of the regulation. FvCaMLP is different from other known EF hand proteins. It shows sequence similarity to both CaMs and myosin regulatory light chain (Cdc4), but has properties typical of a calmodulin, like binding of 45Ca2+, heat stability and Ca2+ dependent electrophoretic shift. Hence, FvCaMLP can be considered a new addition to the category of unconventional Ca2+ binding transcriptional regulators.

Brassinosteroid hormones (BRs) multitask to smoothly regulate a broad spectrum of vital physiological processes in plants, such as cell division, cell expansion, differentiation, seed germination, xylem differentiation, reproductive development and light responses (photomorphogenesis and skotomorphogenesis). Their importance is inferred when visible abnormalities arise in plant phenotypes due to suboptimal or supraoptimal hormone levels. This group of steroidal hormones are major growth regulators, having pleiotropic effects and conferring abiotic stress resistance to plants. Numerous abiotic stresses are the cause of significant loss in agricultural yield globally. However, plants are well equipped with efficient stress combat machinery. Scavenging reactive oxygen species (ROS) is a unique mechanism to combat the deleterious effects of abiotic stresses. In light of numerous reports in the past two decades, the complex BR signaling under different stress conditions (drought, salinity, extreme temperatures and heavy metals/metalloids) that drastically hinders the normal metabolism of plants is gradually being untangled and revealed. Thus, crop improvement has substantial potential by tailoring either the brassinosteroid signaling, biosynthesis pathway or perception. This review aims to explore and dissect the actual mission of BRs in signaling cascades and summarize their positive role with respect to abiotic stress tolerance.