Genetic, Epigenetic, Genomic and Microbial Approaches to Enhance Salt Tolerance of Plants: A Comprehensive Review

Globally, soil salinity has been on the rise owing to various factors that are both human and environmental. The abiotic stress caused by soil salinity has become one of the most damaging abiotic stresses faced by crop plants, resulting in significant yield losses. Salt stress induces physiological and morphological modifications in plants as a result of significant changes in gene expression patterns and signal transduction cascades. In this comprehensive review, with a major focus on recent advances in the field of plant molecular biology, we discuss several approaches to enhance salinity tolerance in plants comprising various classical and advanced genetic and genetic engineering approaches, genomics and genome editing technologies, and plant growth-promoting rhizobacteria (PGPR)-based approaches. Furthermore, based on recent advances in the field of epigenetics, we propose novel approaches to create and exploit heritable genome-wide epigenetic variation in crop plants to enhance salinity tolerance. Specifically, we describe the concepts and the underlying principles of epigenetic recombinant inbred lines (epiRILs) and other epigenetic variants and methods to generate them. The proposed epigenetic approaches also have the potential to create additional genetic variation by modulating meiotic crossover frequency.

Enhancement of Agricultural Crops: A CRISPR/Cas9-Based Approach

Horticultural crops are indispensable agricultural food materials with all essential nutrients. Though, severe threats like pests, diseases, and adverse abiotic factors will affect their productivity and quality. This permits to promote sustainable agriculture by utilizing the recent biotechnological approach to tackle the mentioned issues. In recent year’s genome editing technologies has become one of the most executed genetic tools which altered plant molecular biology. Recently, CRISPR-Cas utilizes for its high target specificity, easier design, and higher success rate. This chapter deals with recent advances in CRISPR/Cas9 technology in horticultural crops in response to the enrichment of essential metabolites, which was achieved by introducing the viral genome to the host via CRISPR-mediated targeted mutation. Furthermore, the strategies based on CRISPR/Cas9 targeted modifications of genes in crop species such as rice, wheat, and soy will be discussed. Finally, we discuss the challenges, improvements, and prospective applications of this cutting-edge technology.

Multiplex Genome-Editing Technologies for Revolutionizing Plant Biology and Crop Improvement

Multiplex genome-editing (MGE) technologies are recently developed versatile bioengineering tools for modifying two or more specific DNA loci in a genome with high precision. These genome-editing tools have greatly increased the feasibility of introducing desired changes at multiple nucleotide levels into a target genome. In particular, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) [CRISPR/Cas] system-based MGE tools allow the simultaneous generation of direct mutations precisely at multiple loci in a gene or multiple genes. MGE is enhancing the field of plant molecular biology and providing capabilities for revolutionizing modern crop-breeding methods as it was virtually impossible to edit genomes so precisely at the single base-pair level with prior genome-editing tools, such as zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs). Recently, researchers have not only started using MGE tools to advance genome-editing applications in certain plant science fields but also have attempted to decipher and answer basic questions related to plant biology. In this review, we discuss the current progress that has been made toward the development and utilization of MGE tools with an emphasis on the improvements in plant biology after the discovery of CRISPR/Cas9. Furthermore, the most recent advancements involving CRISPR/Cas applications for editing multiple loci or genes are described. Finally, insights into the strengths and importance of MGE technology in advancing crop-improvement programs are presented.

Progress in Research on the Mechanisms Underlying Chloroplast-Involved Heat Tolerance in Plants

Global warming is a serious challenge plant production has to face. Heat stress not only affects plant growth and development but also reduces crop yield and quality. Studying the response mechanisms of plants to heat stress will help humans use these mechanisms to improve the heat tolerance of plants, thereby reducing the harm of global warming to plant production. Research on plant heat tolerance has gradually become a hotspot in plant molecular biology research in recent years. In view of the special role of chloroplasts in the response to heat stress in plants, this review is focusing on three perspectives related to chloroplasts and their function in the response of heat stress in plants: the role of chloroplasts in sensing high temperatures, the transmission of heat signals, and the improvement of heat tolerance in plants. We also present our views on the future direction of research on chloroplast related heat tolerance in plants.

Efficient method for isolation of high-quality RNA from Psidium guajava L. tissues

Acquiring high-quality RNA in sufficient amounts is crucial in plant molecular biology and genetic studies. Several methods for RNA extraction from plants are available in the literature, mainly due to the great biochemical diversity present in each species and tissue, which can complicate or prevent the extraction. Psidium guajava (Myrtaceae family) is a perennial fruit tree of medicinal and economic value; nevertheless, only a few molecular studies are available for the species. One reason is the difficulty in obtaining RNA due to the content of the samples, which are rich in polyphenols, polysaccharides, and secondary metabolites. Furthermore, there are few studies available for the isolation of RNA from guava or Psidium samples, which hampers advances in the study of the genus. Here, quality and yields of RNA isolates were compared using six extraction protocols: two protocols based on the application of cetyltrimethylammonium bromide (CTAB) lysis buffer, one protocol which uses the TRIzol reagent, one which applies guanidine thiocyanate lysis buffer followed by organic phase extraction, and two commercial kits (PureLink RNA Mini Kit and RNeasy Plant Mini Kit). The CTAB-based method provided the highest RNA yields and quality for five different tissues (flower bud, immature leaf, young leaf, mature leaf, and root), genotypes, and stress conditions. For the most efficient protocol, the average yield of RNA from guava leaves was 203.06 μg/g of tissue, and the A260/A280 and A260/A230 ratios were 2.1 and 2.2, respectively. RT-qPCR analysis demonstrated that the purity of the samples was sufficient for molecular biology experiments. CTAB-based methods for RNA isolation were found to be the most efficient, providing the highest RNA yields and quality for tissues from P. guajava. Additionally, they were compatible for downstream RNA-based applications, besides being simple and cost-effective.

Genetic and Molecular Factors Determining Grain Weight in Rice

Grain weight is one of the major factors determining single plant yield production of rice and other cereal crops. Research has begun to reveal the regulatory mechanisms underlying grain weight as well as grain size, highlighting the importance of this research for plant molecular biology. The developmental trait of grain weight is affected by multiple molecular and genetic aspects that lead to dynamic changes in cell division, expansion and differentiation. Additionally, several important biological pathways contribute to grain weight, such as ubiquitination, phytohormones, G-proteins, photosynthesis, epigenetic modifications and microRNAs. Our review integrates early and more recent findings, and provides future perspectives for how a more complete understanding of grain weight can optimize strategies for improving yield production. It is surprising that the acquired wealth of knowledge has not revealed more insights into the underlying molecular mechanisms. To accelerating molecular breeding of rice and other cereals is becoming an emergent and critical task for agronomists. Lastly, we highlighted the importance of leveraging gene editing technologies as well as structural studies for future rice breeding applications.

Plant Proteomic Research 3.0: Challenges and Perspectives

Advancements in high-throughput “Omics” techniques have revolutionized plant molecular biology research [...]

Efficient Method for Isolation of High-Quality RNA from Psidium Guajava L. Tissues

Background: Acquiring high-quality RNA in sufficient amounts is necessary in plant molecular biology and genetic studies. Several methods of RNA extraction from plants are available in the literature, mainly due to the great biochemical diversity present in each species and tissue, which can complicate or prevent the extraction. Psidium guajava (Myrtaceae family) is a perennial fruit tree of medicinal and economic value; nevertheless, only few molecular and omics studies are available for the species. One reason for this fact is the difficulty in obtaining the RNA due to the content of the samples, which are rich in polyphenols, polysaccharides and secondary metabolites. Furthermore, there is still no tested or standardized method available for the isolation of RNA from guava or Psidium samples, which hampers advances in the genus.Results: Here we compare the quality and yields of RNA isolated using six extraction protocols: two based on the application of cetyltrimethylammonium bromide (CTAB) lysis buffer, two commercial kits (PureLink™ RNA Mini Kit and RNeasy® Plant Mini Kit), one using the TRIzol® reagent, and one applying guanidine thiocyanate lysis buffer followed by organic phase extraction. RNA integrity, quality and yields were assessed by agarose gel electrophoresis and spectrophotometry. The CTAB-based method provided the highest RNA yields and quality for five different tissues (flower bud, immature leaf, young leaf, mature leaf and root), genotypes and stress conditions. For the most efficient protocol, the average yield of RNA from guava leaf was 210.4 μg/g of tissue, and the A260/A280 and A260/A230 ratios were 2.1 and 2.2, respectively. RT-PCR analysis demonstrated that the purity of the samples was sufficient for molecular biology experiments.Conclusion: CTAB-based methods for RNA isolation were found to be the most efficient, providing the highest RNA yields and quality for tissues from P. guajava. Additionally, they demonstrated to be compatible for downstream RNA-based applications, besides showing the advantages of lower cost and time investments.