Degradome comparison between wild and cultivated rice identifies differential targeting by miRNAs

Background Small non-coding (s)RNAs are involved in the negative regulation of gene expression, playing critical roles in genome integrity, development and metabolic pathways. Targeting of RNAs by ribonucleoprotein complexes of sRNAs bound to Argonaute (AGO) proteins results in cleaved RNAs having precise and predictable 5` ends. While tools to study sliced bits of RNAs to confirm the efficiency of sRNA-mediated regulation are available, they are sub-optimal. In this study, we provide an improvised version of a tool with better efficiency to accurately validate sRNA targets. Results Here, we improvised the CleaveLand tool to identify additional micro (mi)RNA targets that belong to the same family and also other targets within a specified free energy cut-off. These additional targets were otherwise excluded during the default run. We employed these tools to understand the sRNA targeting efficiency in wild and cultivated rice, sequenced degradome from two rice lines, O. nivara and O. sativa indica Pusa Basmati-1 and analyzed variations in sRNA targeting. Our results indicate the existence of multiple miRNA-mediated targeting differences between domesticated and wild species. For example, Os5NG4 was targeted only in wild rice that might be responsible for the poor secondary wall formation when compared to cultivated rice. We also identified differential mRNA targets of secondary sRNAs that were generated after miRNA-mediated cleavage of primary targets. Conclusions We identified many differentially targeted mRNAs between wild and domesticated rice lines. In addition to providing a step-wise guide to generate and analyze degradome datasets, we showed how domestication altered sRNA-mediated cascade silencing during the evolution of indica rice.

Genotype calling and haplotype inference from low coverage sequence data in heterozygous plant genome using HetMap

Many software packages and pipelines had been developed to handle the sequence data of the model species. However, Genotyping from complex heterozygous plant genome needs further improvement on the previous methods. Here we present a new pipeline available at https://github.com/Ncgrhg/HetMapv1) for variant calling and missing genotype imputation from low coverage sequence data for heterozygous plant genomes. To check the performance of the HetMap on the real sequence data, HetMap was applied to both the F1 hybrid rice population which consists of 1495 samples and wild rice population with 446 samples. Four high coverage sequence hybrid rice accessions and two high coverage sequence wild rice accessions, which were also included in low coverage sequence data, are used to validate the genotype inference accuracy. The validation results showed that HetMap archived significant improvement in heterozygous genotype inference accuracy (13.65% for hybrid rice, 26.05% for wild rice) and total accuracy compared with other similar software packages. The application of the new genotype with the genome wide association study also showed improvement of association power in two wild rice phenotypes. It could archive high genotype inference accuracy with low sequence coverage with a small population size with both the natural population and constructed recombination population. HetMap provided a powerful tool for the heterozygous plant genome sequence data analysis, which may help the discover of new phenotype regions for the plant species with complex heterozygous genome.

Chromosome-level genome assembly of Zizania latifolia provides insights into its seed shattering and phytocassane biosynthesis

Chinese wild rice (Zizania latifolia; family: Gramineae) is a valuable medicinal homologous grain in East and Southeast Asia. Here, using Nanopore sequencing and Hi-C scaffolding, we generated a 547.38 Mb chromosome-level genome assembly comprising 332 contigs and 164 scaffolds (contig N50 = 4.48 Mb; scaffold N50 = 32.79 Mb). The genome harbors 38,852 genes, with 52.89% of the genome comprising repetitive sequences. Phylogenetic analyses revealed close relation of Z. latifolia to Leersia perrieri and Oryza species, with a divergence time of 19.7–31.0 million years. Collinearity and transcriptome analyses revealed candidate genes related to seed shattering, providing basic information on abscission layer formation and degradation in Z. latifolia. Moreover, two genomic blocks in the Z. latifolia genome showed good synteny with the rice phytocassane biosynthetic gene cluster. The updated genome will support future studies on the genetic improvement of Chinese wild rice and comparative analyses between Z. latifolia and other plants.

Genome-wide development of lncRNA-derived-SSR markers for Dongxiang wild rice (Oryza rufipogon Griff.)

Dongxiang wild rice (Oryza rufipogon Griff.) (DXWR) is the northernmost distributed wild rice found in the world. Similar to other populations of O. rufipogon, DXWR contains a large number of agronomically valuable genes, which makes it a natural gene pool for rice breeding. Molecular markers, especially simple repeat sequence (SSR) markers, play important roles in plant breeding. Although a large number of SSR markers have been developed, most of them are derived from the genome coding sequences, rarely from non-coding sequences. Meanwhile, long non-coding RNAs (lncRNAs), which are derived from the transcription of non-coding sequences, play vital roles in plant growth, development and stress responses. In our previous study, we obtained 1655 lncRNA transcripts from DXWR using strand-specific RNA sequencing. In this study, 1878 SSR loci were detected from the lncRNA sequences of DXWR, and 1258 lncRNA-derived-SSR markers were developed on the genome-wide scale. To verify the validity and applicability of these markers, 72 pairs of primers were randomly selected to test 44 rice accessions. The results showed that 42 (58.33%) pairs of primers have abundant polymorphism among these rice materials; the polymorphism information content values ranged from 0.04 to 0.87 with an average of 0.50; the genetic diversity index of SSR loci varied from 0.04 to 0.88 with an average of 0.56; and the number of alleles per marker ranged from 2 to 11 with an average of 4.36. Thus, we concluded that these lncRNA-derived-SSR markers are a very useful source for future basic and applied research.

Photosynthetic traits of Australian wild rice (Oryza australiensis) confer tolerance to extreme daytime temperatures

Key message A wild relative of rice from the Australian savannah was compared with cultivated rice, revealing thermotolerance in growth and photosynthetic processes and a more robust carbon economy in extreme heat. Abstract Above ~ 32 °C, impaired photosynthesis compromises the productivity of rice. We compared leaf tissues from heat-tolerant wild rice (Oryza australiensis) with temperate-adapted O. sativa after sustained exposure to heat, as well as diurnal heat shock. Leaf elongation and shoot biomass in O. australiensis were unimpaired at 45 °C, and soluble sugar concentrations trebled during 10 h of a 45 °C shock treatment. By contrast, 45 °C slowed growth strongly in O. sativa. Chloroplastic CO2 concentrations eliminated CO2 supply to chloroplasts as the basis of differential heat tolerance. This directed our attention to carboxylation and the abundance of the heat-sensitive chaperone Rubisco activase (Rca) in each species. Surprisingly, O. australiensis leaves at 45 °C had 50% less Rca per unit Rubisco, even though CO2 assimilation was faster than at 30 °C. By contrast, Rca per unit Rubisco doubled in O. sativa at 45 °C while CO2 assimilation was slower, reflecting its inferior Rca thermostability. Plants grown at 45 °C were simultaneously exposed to 700 ppm CO2 to enhance the CO2 supply to Rubisco. Growth at 45 °C responded to CO2 enrichment in O. australiensis but not O. sativa, reflecting more robust carboxylation capacity and thermal tolerance in the wild rice relative.

Identification of Phosphorus Stress Related Proteins in the Seedlings of Dongxiang Wild Rice (Oryza Rufipogon Griff.) Using Label-Free Quantitative Proteomic Analysis

Phosphorus (P) deficiency tolerance in rice is a complex character controlled by polygenes. Through proteomics analysis, we could find more low P tolerance related proteins in unique P-deficiency tolerance germplasm Dongxiang wild rice (Oryza Rufipogon, DXWR), which will provide the basis for the research of its regulation mechanism. In this study, a proteomic approach as well as joint analysis with transcriptome data were conducted to identify potential unique low P response genes in DXWR during seedlings. The results showed that 3589 significant differential accumulation proteins were identified between the low P and the normal P treated root samples of DXWR. The degree of change was more than 1.5 times, including 60 up-regulated and 15 downregulated proteins, 24 of which also detected expression changes of more than 1.5-fold in the transcriptome data. Through quantitative trait locus (QTLs) matching analysis, seven genes corresponding to the significantly different expression proteins identified in this study were found to be uncharacterized and distributed in the QTLs interval related to low P tolerance, two of which (LOC_Os12g09620 and LOC_Os03g40670) were detected at both transcriptome and proteome levels. Based on the comprehensive analysis, it was found that DXWR could increase the expression of purple acid phosphatases (PAPs), membrane location of P transporters (PTs), rhizosphere area, and alternative splicing, and it could decrease reactive oxygen species (ROS) activity to deal with low P stress. This study would provide some useful insights in cloning the P-deficiency tolerance genes from wild rice, as well as elucidating the molecular mechanism of low P resistance in DXWR.

Commercial Cultivation of Australian Wild Oryza spp.: A Review and Conceptual Framework for Future Research Needs

Wild Oryza species are being targeted for commercial cultivation due to their high nutritional grain profile, and their association with Aboriginal people in many regions. Australian wild Oryza species have potential as high-value, low-volume, culturally identified, and nutritious food, especially in gourmet food, tourism, restaurants, and value-added products. However, the basic agronomic protocols for their cultivation as a field crop are unknown. In this review, we identify the major factors supporting the commercial production of wild Oryza, including their stress-tolerant capacity, excellent grain quality attributes, and Indigenous cultural identification of their grains. The key challenges to be faced during the development of a wild rice industry are also discussed which include management barriers, processing issues, undesirable wild traits, and environmental concern. This manuscript proposes the use of agronomic research, in combination with breeding programs, as an overarching framework for the conceptualization and implementation of a successful wild rice industry, using the North American wild rice industry as a case study. The framework also suggests an integrated system that connects producers, industry, and government stakeholders. The suggested procedures for developing a wild rice industry in Australia are also applicable for other wild Oryza species.

Characterization of Wild Rice - Oryza Species Complexes in Sri Lanka

Rice is the staple food crop in Sri Lanka, which occupies 34% (0.77/million ha) of the total cultivated area. Sri Lanka currently produces 2.7 million tonnes of rough rice annually and satisfies around 95% of the domestic requirement. In Sri Lanka, genus Oryza consists of two species complexes, O. sativa (AA) and O. officinalis (CC). These two complexes are both pan tropical and have very similar overall distribution. Five wild rice species are reported in Sri Lanka, (O. nivara [AA], O. rufipogan (AA) O. eichengeri [CC], O. rhizomatis (CC) and O. granulate (GG). O. rhizomatis has been reported only in Sri Lanka and considered endemic to Sri Lanka. Recent studies demonstrated, the reliance on single source of information could mislead results in the phylogenetic inferences due to analytical inconsistency and biological processes. Therefore, exact number of wild rice species in Sri Lanka becomes uncertain and the necessity arises to assess Oryza species complexes in Sri Lanka using morphological, anatomical, and molecular information to enumerate number of species within each Oryza complex and characterization of species and species complexes. The study revealed, characterization of wild rice species, to a certain extent, can be made through morphological and anatomical characters, specially lamina anatomical characters. Molecular information is more reliable in delimitation of wild rice species complexes in Sri Lanka. O. rhizomatis and O. eichingeri (CC) are well separated from the rest of wild rice species (AA). Molecular data revealed, O. nivara and O. rufipogon have undergone independent evolution within Sri Lanka. Well separated five wild rice species are existing in Sri Lanka. Studies on ecological resilience of morphological, anatomical, and molecular studies are very useful for species enumeration of wild rice complexes in Sri Lanka. The findings led to conclude that wild rice species in Sri Lanka are “ecological swarms” and represents allopatric or sympatric populations. A comprehensive knowledge on genetic diversity and population structure of wild rice germplasm in Sri Lanka provides useful information to include these locally adapted and evolved wild rice species in rice crop improvement/breeding.