Genetic mapping of regions associated with root system architecture in rice using MutMap QTL-seq.

The root system architecture is an important complex trait in rice. With changing climatic conditions and soil nutrient deficiencies, there is an immediate need to breed nutrient-use-efficient rice varieties with robust root system architectural (RSA) traits. To map the genomic regions associated with crucial component traits of RSA viz. root length and root volume, a biparental F2 mapping population was developed using TI-128, an Ethyl Methane Sulphonate (EMS) mutant of a mega variety BPT-5204 having high root length (RL) and root volume (RV) with wild type BPT-5204. Extreme bulks having high RL and RV and low RL and RV were the whole genome re-sequenced along with parents. Genetic mapping using the MutMap QTL-Seq approach elucidated two genomic intervals on Chr.12 (3.14-3.74 Mb, 18.11-20.85 Mb), and on Chr.2 (23.18-23.68 Mb) as potential regions associated with both RL and RV. The Kompetitive Allele Specific PCR (KASP) assays for SNPs with delta SNP index near 1 were associated with higher RL and RV in the panel of sixty-two genotypes varying in root length and volume. The KASP_SNPs viz. Chr12_S4 (C→T; Chr12:3243938), located in the 3' UTR region of LOC_Os12g06670 encoding a protein kinase domain-containing protein and Chr2_S6 (C→T; Chr2:23181622) present upstream in the regulator of chromosomal condensation protein LOC_Os2g38350. Validation of these genes using qRT-PCR and in-silico studies using various online tools and databases revealed higher expression in TI-128 as compared to BPT- 5204 at the seedling and panicle initiation stages implying the functional role in enhancing RL and RV.

Impairment in a member of AP2/ERF and F-box family protein enhances complete panicle exsertion in rice.

Complete panicle exsertion (CPE) is an economically important quantitative trait that contributes to grain yield in rice. We deployed an integrated approach for understanding the molecular mechanism of CPE using a stable ethyl methanesulfonate mutant line, CPE-109 of the Samba Mahsuri (SM) variety of rice (Oryza sativa), which exhibits CPE. Two consistent genomic regions were identified for CPE through quantitative trait locus (QTL) mapping [qCPE-4 (28.24-31.22 Mb) and qCPE-12 (2.30-3.18 Mb)] and QTL-sequencing [chr 4 (31.21-33.69 Mb) and chr 12 (0.12-3.15 Mb)]. Two non-synonymous single nucleotide polymorphisms, namely KASP 12-12 (T→C; chr12:1269983) in Os12g0126300, encoding an AP2/ERF transcription factor, and KASP 12-16 (G→A; chr12:1515198) in Os12g0131400, encoding an F-box domain-containing protein, explained 81.05% and 59.61% of the phenotypic variance, respectively, and exhibited strong co-segregation with CPE in F2 mapping populations, advanced generation lines, and CPE-exhibiting SM mutants through KASP assays. Down-regulation of these genes in CPE-109 compared with SM (wild type) was observed in transcriptome sequencing of flag leaves, which was validated through qRT-PCR. We propose that the abrogation of Os12g0126300 and Os12g0131400 in CPE-109 combinatorially influences down-regulation of ethylene biosynthetic genes, Os01g0192900 (ACC synthase) and Os05g0497300 (ethylene-responsive factor-2), and up-regulation of a gibberellic acid synthetic gene, Os06g0569900 (ent-kaurene synthase) and the two cytokinin biosynthetic genes Os07g0486700 (cytokinin-O-glucosyltransferase 2) and Os10g0479500 (similar to carboxy-lyase), which results in complete panicle exsertion.

Multiomics-assisted characterization of rice-Yellow Stem Borer interaction provides genomic and mechanistic insights into stem borer resistance in rice.

KEY MESSAGE: By deploying a multi-omics approach, we unraveled the mechanisms that might help rice to combat Yellow Stem Borer infestation, thus providing insights and scope for developing YSB resistant rice varieties. Yellow Stem Borer (YSB), Scirpophaga incertulas (Walker) (Lepidoptera: Crambidae), is a major pest of rice, that can lead to 20-60% loss in rice production. Effective management of YSB infestation is challenged by the non-availability of adequate sources of resistance and poor understanding of resistance mechanisms, thus necessitating studies for generating resources to breed YSB resistant rice and to understand rice-YSB interaction. In this study, by using bulk-segregant analysis in combination with next-generation sequencing, Quantitative Trait Loci (QTL) intervals in five rice chromosomes were mapped that could be associated with YSB resistance at the vegetative phase in a resistant rice line named SM92. Further, multiple SNP markers that showed significant association with YSB resistance in rice chromosomes 1, 5, 10, and 12 were developed. RNA-sequencing of the susceptible and resistant lines revealed several genes present in the candidate QTL intervals to be differentially regulated upon YSB infestation. Comparative transcriptome analysis revealed a putative candidate gene that was predicted to encode an alpha-amylase inhibitor. Analysis of the transcriptome and metabolite profiles further revealed a possible link between phenylpropanoid metabolism and YSB resistance. Taken together, our study provides deeper insights into rice-YSB interaction and enhances the understanding of YSB resistance mechanism. Importantly, a promising breeding line and markers for YSB resistance have been developed that can potentially aid in marker-assisted breeding of YSB resistance among elite rice cultivars.

Strong culm: a crucial trait for developing next-generation climate-resilient rice lines.

Lodging, a phenomenon characterized by the bending or breaking of rice plants, poses substantial constraints on productivity, particularly during the harvesting phase in regions susceptible to strong winds. The rice strong culm trait is influenced by the intricate interplay of genetic, physiological, epigenetic, and environmental factors. Stem architecture, encompassing morphological and anatomical attributes, alongside the composition of both structural and non-structural carbohydrates, emerges as a critical determinant of lodging resistance. The adaptive response of the rice culm to various biotic and abiotic environmental factors further modulates the propensity for lodging. Advancements in next-generation sequencing technologies have expedited the genetic dissection of lodging resistance, enabling the identification of pertinent genes, quantitative trait loci, and novel alleles. Concurrently, contemporary breeding strategies, ranging from biparental approaches to more sophisticated methods such as multi-parent-based breeding, gene pyramiding, genomic selection, genome-wide association studies, and haplotype-based breeding, offer perspectives on the genetic underpinnings of culm strength. This review comprehensively delves into physiological attributes, culm histology, epigenetic determinants, and gene expression profiles associated with lodging resistance, with a specialized focus on leveraging next-generation sequencing for candidate gene discovery.

Long non-coding RNA-mediated epigenetic response for abiotic stress tolerance in plants.

Plants perceive environmental fluctuations as stress and confront several stresses throughout their life cycle individually or in combination. Plants have evolved their sensing and signaling mechanisms to perceive and respond to a variety of stresses. Epigenetic regulation plays a critical role in the regulation of genes, spatiotemporal expression of genes under stress conditions and imparts a stress memory to encounter future stress responses. It is quintessential to integrate our understanding of genetics and epigenetics to maintain plant fitness, achieve desired genetic gains with no trade-offs, and durable long-term stress tolerance. The long non-coding RNA >200 nts having no coding potential (or very low) play several roles in epigenetic memory, contributing to the regulation of gene expression and the maintenance of cellular identity which include chromatin remodeling, imprinting (dosage compensation), stable silencing, facilitating nuclear organization, regulation of enhancer-promoter interactions, response to environmental signals and epigenetic switching. The lncRNAs are involved in a myriad of stress responses by activation or repression of target genes and hence are potential candidates for deploying in climate-resilient breeding programs. This review puts forward the significant roles of long non-coding RNA as an epigenetic response during abiotic stresses in plants and the prospects of deploying lncRNAs for designing climate-resilient plants.

Genome-wide association studies in rice germplasm reveal significant genomic regions for root and yield-related traits under aerobic and irrigated conditions.

The development of nutrient-use efficient rice lines is a priority amidst the changing climate and depleting resources viz., water, land, and labor for achieving sustainability in rice cultivation. Along with the traditional transplanted irrigated system of cultivation, the dry direct-seeded aerobic system is gaining ground nationwide. The root-related traits play a crucial role in nutrient acquisition, adaptation and need to be concentrated along with the yield-attributing traits. We phenotyped an association panel of 118 rice lines for seedling vigour index (SVI) traits at 14 and 21 days after sowing (DAS), root-related traits at panicle initiation (PI) stage in polythene bags under controlled aerobic condition, yield and yield-related traits under the irrigated condition at ICAR-IIRR, Hyderabad, Telangana; irrigated and aerobic conditions at ARS, Dhadesugur, Raichur, Karnataka. The panel was genotyped using simple sequence repeats (SSR) markers and genome-wide association studies were conducted for identifying marker-trait associations (MTAs). Significant correlations were recorded for root length, root dry weight with SVI, root volume at the PI stage, number of productive tillers per plant, spikelet fertility, the total number of grains per panicle with grain yield per plant under irrigated conditions, and the total number of grains per panicle with grain yield per plant under aerobic condition. The panel was divided into three sub-groups (K = 3) and correlated with the principal component analysis. The maximum number of MTAs were found on chromosomes 2, 3, and 12 with considerable phenotypic variability. Consistent MTAs were recorded for SVI traits at 14 and 21 DAS (RM25310, RM80, RM22961, RM1385), yield traits under irrigated conditions (RM2584, RM5179, RM410, RM20698, RM14753) across years at ICAR-IIRR, grain yield per plant (RM22961, RM1146) under the aerobic condition, grain yield per plant at irrigated ICAR-IIRR and SVI (RM5501), root traits at PI stage (RM2584, RM80, RM410, RM1146, RM18472). Functionally relevant genes near the MTAs through in-silico expression analysis in root and panicle tissues viz., HBF2 bZIP transcription factor, WD40 repeat-like domain, OsPILS6a auxin efflux carrier, WRKY108, OsSCP42, OsMADS80, nodulin-like domain-containing protein, amino acid transporter using various rice expression databases were identified. The identified MTAs and rice lines having high SVI traits (Langphou, TI-128, Mouli, TI-124, JBB-631-1), high yield under aerobic (Phouren, NPK-43, JBB-684, Ratnamudi, TI-112), irrigated conditions (KR-209, KR-262, Phouren, Keibi-Phou, TI-17), robust root traits like root length (MoirangPhou-Angouba, Wangoo-Phou, JBB-661, Dissi, NPK-45), root volume (Ratnachudi, KJ-221, Mow, Heimang-Phou, PUP-229) can be further employed in breeding programs for the targeted environments aimed at improving seedling vigour, yield-related traits under irrigated condition, aerobic condition as adaptability to water-saving technology.

Combined strategy employing MutMap and RNA-seq reveals genomic regions and genes associated with complete panicle exsertion in rice.

Complete panicle exsertion (CPE) in rice is an important determinant of yield and a desirable trait in breeding. However, the genetic basis of CPE in rice still remains to be completely characterized. An ethyl methane sulfonate (EMS) mutant line of an elite cultivar Samba Mahsuri (BPT 5204), displaying stable and consistent CPE, was identified and named as CPE-110. MutMap and RNA-seq were deployed for unraveling the genomic regions, genes, and markers associated with CPE. Two major genomic intervals, on chromosome 8 (25668481-25750456) and on chromosome 11 (20147154-20190400), were identified to be linked to CPE through MutMap. A non-synonymous SNP (G/A; Chr8:25683828) in the gene LOC_Os08g40570 encoding pyridoxamine 5'-phosphate oxidase with the SNP index 1 was converted to Kompetitive allele-specific PCR (KASP) marker. This SNP (KASP 8-1) exhibited significant association with CPE and further validated through assay in the F2 mapping population, released varieties and CPE exhibiting BPT 5204 mutant lines. RNA-seq of the flag leaves at the booting stage, 1100 genes were upregulated and 1305 downregulated differentially in CPE-110 and BPT 5204. Metabolic pathway analysis indicated an enrichment of genes involved in photosynthesis, glyoxylate, dicarboxylate, porphyrin, pyruvate, chlorophyll, carotenoid, and carbon metabolism. Further molecular and functional studies of the candidate genes could reveal the mechanistic aspects of CPE. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01412-1.

Current technological interventions and applications of CRISPR/Cas for crop improvement.

Efficient and innovative breeding strategies are immensely required to meet the global food demand, nutritional security and sustainable agriculture. Genome editing tools have emerged as an effective technology for site-directed genome modification causing the change in gene expression and protein function for the improvement of various important traits in particular the CRISPR/Cas (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated protein). As the technology evolved with time, advances have been observed like prime editing, base editing, PAMless editing, Drosha based editing with multiple targets having the potential to fulfill the regulatory processes around the world. These recent interventions are highly proficient, cost-efficient, user-friendly, and holds promise for a major revolution in basic and applied plant biology research in the ever-evolving climatic conditions. In the review, we have discussed the most recent technologies and advances for CRISPR/Cas editing in plants.