Molecular Approaches for Insect Pest Management in Rice

AbstractThis chapter focuses on the progress made in using molecular tools in understanding resistance in rice to insect pests and breeding rice for multiple and durable insect resistance. Currently, molecular markers are being extensively used to tag, map, introgress, and clone plant resistance genes against gall midge, planthoppers, and leafhoppers. Studies on cloned insect resistance genes are leading to a better understanding of plant defense against insect pests under different feeding guilds. While marker-assisted breeding is successfully tackling problems in durable and multiple pest resistance in rice, genomics of plants and insects has identified RNAi-based gene silencing as an alternative approach for conferring insect resistance. The use of these techniques in rice is in the developmental stage, with the main focus on brown planthopper and yellow stem borer. CRISPR-based genome editing techniques for pest control in plants has just begun. Insect susceptibility genes (negative regulators of resistance genes) in plants are apt targets for this approach while gene drive in insect populations, as a tool to study rice-pest interactions, is another concept being tested. Transformation of crop plants with diverse insecticidal genes is a proven technology with potential for commercial success. Despite advances in the development and testing of transgenic rice for insect resistance, no insect-resistant rice cultivar is now being commercially cultivated. An array of molecular tools is being used to study insect-rice interactions at transcriptome, proteome, metabolome, mitogenome, and metagenome levels, especially with reference to BPH and gall midge, and such studies are uncovering new approaches for insect pest management and for understanding population genetics and phylogeography of rice pests. Thus, it is evident that the new knowledge being gained through these studies has provided us with new tools and information for facing future challenges. However, what is also evident is that our attempts to manage rice pests cannot be a one-time effort but must be a continuing one

Integrated Physiological, Biochemical, and Molecular Analysis Identifies Important Traits and Mechanisms Associated with Differential Response of Rice Genotypes to Elevated Temperature

In changing climate, heat stress caused by high temperature poses a serious threat to rice cultivation. A multiple organizational analysis at physiological, biochemical and molecular level is required to fully understand the impact of elevated temperature in rice. This study was aimed at deciphering the elevated temperature response in eleven popular and mega rice cultivars widely grown in India. Physiological and biochemical traits specifically membrane thermostability (MTS), antioxidants, and photosynthesis were studied at vegetative and reproductive phases which were used to establish a correlation with grain yield under stress. Several useful traits in different genotypes were identified which will be important resource to develop high temperature tolerant rice cultivars. Interestingly, Nagina22 emerged as best performer in terms of yield as well as expression of physiological and biochemical traits at elevated temperature. It showed lesser relative injury, lesser reduction in chlorophyll content, increased super oxide dismutase, catalase and peroxidase activity, lesser reduction in net photosynthetic rate (PN), high transpiration rate (E) and other photosynthetic/ fluorescence parameters contributing to least reduction in spikelet fertility and grain yield at elevated temperature. Further, expression of 14 genes including heat shock transcription factors and heat shock proteins was analyzed in Nagina22 (tolerant) and Vandana (susceptible) at flowering phase, strengthening the fact that N22 performs better at molecular level also during elevated temperature. This study shows that elevated temperature response is complex and involves multiple biological processes which are needed to be characterized to address the challenges of future climate extreme conditions

Fine mapping and sequence analysis reveal a promising candidate gene encoding a novel NB-ARC domain derived from wild rice (Oryza officinalis) that confers bacterial blight resistance

Bacterial blight disease of rice caused by Xanthomonas oryzae pv. oryzae (Xoo) is one of the most serious constraints in rice production. The most sustainable strategy to combat the disease is the deployment of host plant resistance. Earlier, we identified an introgression line, IR 75084-15-3-B-B, derived from Oryza officinalis possessing broad-spectrum resistance against Xoo. In order to understand the inheritance of resistance in the O. officinalis accession and identify genomic region(s) associated with resistance, a recombinant inbred line (RIL) mapping population was developed from the cross Samba Mahsuri (susceptible to bacterial blight) × IR 75084-15-3-B-B (resistant to bacterial blight). The F2 population derived from the cross segregated in a phenotypic ratio of 3: 1 (resistant susceptible) implying that resistance in IR 75084-15-3-B-B is controlled by a single dominant gene/quantitative trait locus (QTL). In the F7 generation, a set of 47 homozygous resistant lines and 47 homozygous susceptible lines was used to study the association between phenotypic data obtained through screening with Xoo and genotypic data obtained through analysis of 7K rice single-nucleotide polymorphism (SNP) chip. Through composite interval mapping, a major locus was detected in the midst of two flanking SNP markers, viz., Chr11.27817978 and Chr11.27994133, on chromosome 11L with a logarithm of the odds (LOD) score of 10.21 and 35.93% of phenotypic variation, and the locus has been named Xa48t. In silico search in the genomic region between the two markers flanking Xa48t identified 10 putatively expressed genes located in the region of interest. The quantitative expression and DNA sequence analysis of these genes from contrasting parents identified the Os11g0687900 encoding an NB-ARC domain-containing protein as the most promising gene associated with resistance. Interestingly, a 16-bp insertion was noticed in the untranslated region (UTR) of the gene in the resistant parent, IR 75084-15-3-B-B, which was absent in Samba Mahsuri. The association of Os11g0687900 with resistance phenotype was further established by sequence-based DNA marker analysis in the RIL population. A co-segregating PCR-based INDEL marker, Marker_Xa48, has been developed for use in the marker-assisted breeding of Xa48t

Cloning and characterization of drought responsive partial gene sequence(s) from Oryza sativa L. subsp. Indica

387-392Differential display gels were run for the drought tolerant (N-22) and drought susceptible (Panidhan) genotypes of rice (Oryza sativa) to identify the genes showing differential expression with respect to moisture stress. Differential cDNA products were cloned in PCR-Trap vector and analyzed for differential expression by Northern hybridization. Two clones namely R4A and R7G were found to be associated with water deficit stress (WDS). Sequencing revealed an insert of 244 bp in the clone R4A. BLASTN and FASTA results showed that R4A had maximum homology with a full-length cDNA clone: 002-110-H10 and OSJNBa006109.12 protein. GO classification suggested that it had β-glucosidase motif which had been implicated in ABA mobilization and thereby ABA dependent gene expression

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Not AvailableVirus–host interaction at cellular and intra-cellular level is a constantly evolving process which results either in the host to resist or the virus to break host immunity and to establish the disease. We have put extensive efforts to understand the genomic organization and gene functions of important viral proteins involved in resisting or avoiding host antiviral responses mediated by RNA interference or Ubiquitin–Proteasome Pathway. Nearly two decades of dedicated research on three agriculturally important viruses revealed genetic and epigenetic regulation of host to induce the defense/immunity responses. The microRNA and auxin regulated development of disease symptoms and the role of temperature in optimizing the interactions of viral protein with host small RNAs are intriguing observations. Owing to the complexity of the dynamic interactions between plant and virus, this research field will always be challenging and fascinating.Not Availabl

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Not AvailableHigh-temperature stress is a major abiotic stress that affects various biological processes Of plants Such as biochemical and physiological response, growth, development, and yield. High-temperature Stress has critical effects at cellular and molecular levels also. The increased concentration of regulatory proteins such as heat shock transcription factors (Hsfs) is a major molecular response that occurs during heat stress. These regulatory proteins in turn regulate the expression of heat shock protein (HSP) genes that act as critical players during stress to maintain cell homeostasis. Besides HSPs, the other metabolic and regulatory genes, signalling compounds, compatible osmolytes, and antioxidants too play an important role during Heat stress in plants. Apart From the protein-coding genes, recent studies have shown that noncoding microns (miRNAs) Also play a key role during heat stress by modulating the gene expression at the transcription and post-transcriptional evel. The transcriptome approaches are important to understand the molecular and cellular changes occurring in response to heat stress. The approaches rely mostly by adopting the traditional methods like Northern blot/RNA blot and reverse transcription PCR (RT-PCR), Where the expression of the genes can be studied in different tissues and cells, whereas the extent of their expression can be achieved by quantitative PCR Or real time PCR. Further, The genome-wide expression profiling tools such as microarray analysis, next-generation sequencing, and RNA Sequencing offer a great potential in this direction. This chapter primarily provides the current understanding on the role of regulatory genes (transcription factors), HSP genes, metabolic genes, signaling compounds, osmolytes, reactive oxygen species, and miRNAs as well as other small RNAs of plants under high temperature. In addition, it gives a brief account of various transcriptome approaches to study the expression profiling of genes during heat stress.Not Availabl

Key Enzymes and Proteins of Crop Insects as Candidate for RNAi Based Gene Silencing

RNA interference (RNAi) is a mechanism of homology dependent gene silencing present in plants and animals. It operates through 21-24 nucleotides small RNAs which are processed through a set of core enzymatic machinery that involves Dicer and Argonaute proteins. In recent past, the technology has been well appreciated towards the control of plant pathogens and insects through suppression of key genes/proteins of infecting organisms. The genes encoding key enzymes/proteins with the great potential for developing an effective insect control by RNAi approach are actylcholinesterase, cytochrome P450 enzymes, amino peptidase N, allatostatin, allatotropin, tryptophan oxygenase, arginine kinase, vacuolar ATPase, chitin synthase, glutathione-S-transferase, catalase, trehalose phosphate synthase, vitellogenin, hydroxy-3-methylglutaryl coenzyme A reductase, and hormone receptor genes. Through various studies, it is demonstrated that RNAi is a reliable molecular tool which offers great promises in meeting the challenges imposed by crop insects with careful selection of key enzymes/proteins. Utilization of RNAi tool to target some of these key proteins of crop insects through various approaches is described here. The major challenges of RNAi based insect control such as identifying potential targets, delivery methods of silencing trigger, off target effects, and complexity of insect biology are very well illustrated. Further, required efforts to address these challenges are also discussed

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Not AvailableMicroRNAs (miRNAs) are small non-coding RNAs which play an important role in regulating the genes involved in plant growth and development. Several studies showed that miRNAs are involved in plants response to different kinds of abiotic stresses also. In our previous study, temperature responsive miRNAs were predicted in O.sativa. 27 miRNAs were predicted to be novel in rice using homology search. In continuation to our previous study, expression of 14 novel miRNAs was done in shoot and root of 13 days old seedlings of five different rice cultivars using real time PCR. Expression these miRNAs was analyzed in control and high temperature stress environment. Out of 14 predicted novel miRNAs, two novel miRNAs- miR157a and miR165a showed expression in all five rice cultivars. Interestingly, miR165a showed a differential expression pattern among heat tolerant (N22, IR64 and Rasi) and susceptible (Vandana and Sampada) cultivars suggesting that it might have specific role in high temperature tolerance.Not Availabl

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Not AvailableComputational prediction of potential miRNAs and their target genes was performed to identify the miRNAs and genes associated with temperature response in rice. The data of temperatureresponsive miRNAs of Arabidopsis, and miRNAs and whole genome data of rice were used to predict potential miRNAs in O. sativa involved in temperature response. A total of 55 miRNAs were common in both the species. A total of 27 miRNAs were predicted at the first time in rice. Target genes were searched for these 27 miRNAs in rice genome following stringent criteria. Real time PCR based on expression analysis of nine miRNAs showed that majority of the miRNAs were down regulated under heat stress for rice cultivar Nagina 22. Furthermore, miR169, miR1884 and miR160 showed differential expression in root and shoot tissues of rice.Identification and expression studies of miRNAs duringheat stress will advance the understanding of gene regulation under stress in rice.Not Availabl