2016. Adaptation to drought in rice. Dissecting the role of Jamonates for the response to drought in rice

2016.  Adaptation to drought in rice. Dissecting the role of Jamonates for the response to drought in rice

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ABTRACT

The various adaptive responses of drought stressed plants are strongly influenced by chemical messengers called phytohormones, which help the plants to regulate growth and development also in adverse conditions such as drought. One such important phytohormone is jasmonic acid (JA). However, little is known about its direct involvement in drought response or about its cross-talk to other phytohormones such as abscisic acid (ABA), which is intricately involved in drought stress adaptations. Previously, it was shown that the rice mutant cpm2, impaired in the function of allene oxide cyclase (AOC), a key enzyme in JA biosynthesis, was less sensitive to salt stress. In the present study, comparative changes under drought in some physiological traits and in the root proteome of cpm2 and the wild type (WT) were analyzed. When the stomatal conductance in WT and cpm2 under drought conAdition was measured, cpm2 had lower stomatal conductance under drought stress as compared to WT and this also correlated with increased ABA levels in shoots. Under drought, higher Water Use Efficiency (WUE) in cpm2 as compared to WT also indicated improved drought tolerance. Importantly, roots of cpm2 were better developed under both control and moderate drought stress. To assess if the roots of cpm2 and WT respond differentially to drought at the molecular level, root proteome analysis was undertaken using Tandem Mass Tag (TMT) approach. At the protein level, AOC was unique to WT and highly abundant under drought. This confirmed the lack of AOC in cpm2. Another protein, OPDA reductase (0PR7) which is downstream of AOC in JA biosynthesis pathway became more abundant in cpm2 while its amount decreased in the WT in drought stress. These results suggest that OPDA, a precursor of JA, and not JA itself is accumulating in response to drought stress. Analysis of other differentially expressed proteins revealed more active ROS detoxification and metabolism in cpm2 under drought. In addition, a number of proteins involved in pathways related to cell wall remodeling and cell growth were also abundant in cpm2 roots. These results suggested that JA signaling might negatively influence drought tolerance by orchestrating a block on critical morpho-physiological and molecular changes necessary for stress adaptation. Mutant analysis suggested benefits of blocking JA synthesis which might be useful for improving drought tolerance in rice  .

2012. Genome wide association mapping of drought resistance traits in rice

2012.  Genome wide association mapping of drought resistance traits in rice

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ABTRACT

Rice, (Oryza sativa L.) is one of the most important crops providing staple food for a large segment of the world population. Drought stress is a major limitation to rice yields and its stability in rainfed areas. Developing drought resistant cultivars will help to increase production of rice in rainfed area. However, conventional breeding for drought resistance is slow in attaining progress due to poor understanding of genetic control of drought resistance and difficulty in phenotyping breeding progenies for drought tolerance. Molecular markers help in identification of quantitative trait loci (QTLs) associated with drought resistance traits and their use in breeding high yielding rice varieties suitable for drought-prone areas through marker-assisted breeding, thus reducing the need for extensive field testing over space and time. Conventional QTL mapping requires genotyping large mapping progenies derived from biparental crossing, demanding time and labour. Genome-wide association mapping is a powerful approach to identify associations between traits of interest and genetic markers using diverse genotypes. Thus, in the present study, a total of 48 rice accessions from different geographic locations and ecotypes were evaluated for plant production and root traits under irrigated and rainfed conditions in target populations of environment. Rice accessions showed significant differences for plant phenology, production and root traits. The rice accessions were screened using a total of 510 SSR markers selected from across the rice genome. Among them, 323 SSR markers were found to be polymorphic among the accessions. Structure analysis grouped the 48 accessions into six populations, broadly corresponding to the major geographic regions. Genome-wide association analysis using TASSEL identified a total of 444 marker-trait associations at P<0.05, P<0.001 and PO.OOOl for the various traits. Among them, 155 markers were associated with yield traits and 82 markers for root traits in these rice accessions. The markers viz., RM170 chromosome 6 (ch. 6), RM5720 (ch. 7), RM318 (ch. 2), RM3843 (ch. 4), RM29 (ch. 2), RM540 (ch. 6), RM585 (ch. 6) and RM36 (ch. 3) are linked to both yield and root traits under drought stress in this study. These markers were earlier identified to be linked to drought resistance and yield traits in rice through conventional QTL mapping efforts. The consistent markers for yield and root traits can be used in marker-assisted breeding for rainfed rice improvement. Thus, genome wide association mapping is ideal to detect markers linked to plant production and drought resistance traits in rice using diverse genotypes  .

2011. Defense response signaling for disease resistance in rice

2011. Defense response signaling for disease resistance in rice

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ABTRACT

Plant disease resistance is often associated with a type of programmed cell death (PCD) called the hypersensitive response (HR). Upon recognition of pathogen proteins or their activity, the affected cell and surrounding cells commit to the HR to limit pathogen spread throughout the plant. This mechanism of plant disease resistance renders a pathogen avirulent on the host plant. Lesion mimics (LM) are a class of mutant or transgenic plants that spontaneously show lesions resembling the HR in the absence of biotic stress. Based on the association of the LM phenotype to cell death and its similarity to disease symptoms and the HR, this phenotype is a useful tool to dissect and understand the plant defense response.

To identify genes that when mutated result in the LM phenotype in rice, we used a microarray approach. By hybridizing labeled genomic DNA from an allelic series of deletion mutants to an oligonucleotide microarray, we identified candidate genes and genic regions that were deleted in a set of mutants. For one mutant, spll, mutations in a cytochrome P450 gene were confirmed to confer the LM phenotype. A genome browser developed to handle these microarray data is a community resource that enables researchers to rapidly identify untagged deletion mutations in rice.

Members of the 14-3-3 protein family were recently shown to be positive regulators of cell death and the HR in Arabidopsis. In contrast, the work herein shows that a rice 14-3-3 protein is a negative regulator of cell death and resistance. Transgenic plants carrying a construct that silences the rice 14-3-3 gene GF14e exhibit a LM phenotype and enhanced resistance to two distinct rice pathogens, Xanthomonas oryzae pv. oryzae and Rhizoctonia solani. These Gf74<?-silenced plants also showed enhanced expression of genes associated with salicylic acid (SA) mediated defense responses, including members of the peroxidase gene family. The GF/4<?-silenced plants did not show enhanced expression of marker genes associated with the ethylene response pathway, indicating that GF14e may negatively regulate SA mediated defense responses, but does not affect ethylene regulated responses.

Silencing GF14e results in the upregulation of several defense responsive peroxidases. cDNA from GF14e silenced plants was used in quantitative PCR (qPCR) to assay expression of four peroxidase genes. Of these, three showed significant upregulation in 2 weeks after sowing (WAS) and 5 WAS GF1^-silenced plants. The promoters of the three upregulated genes (PO-C1, PoxS.l, Pox22.3) contain at least one W-box element. In contrast, the peroxidase (Pox5.1) that did not show upregulation and is not upregulated in R gene mediated responses had no W-box elements. W-box elements are binding sites for the WRKY class of transcription factors. This result, coupled with bioinformatic predictions of potential rice 14-3-3 clients, and the observation that some WRKY genes are upregulated in GF14e silenced lines indicates that GF14e may negatively regulate WRKY transcription factors related to cell death and defense responses.

Based on the implication that one 14-3-3 protein negatively regulates defense responses related cell death, provides a framework to develop a model for how this protein might function in the plant disease  resistance repsonse  .

2010. Genome organization and genetic diversity of wildrice (Zizania palustris L.)

2010. Genome organization and genetic diversity of wildrice (Zizania palustris L.)

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ABSTRACT

The large amount of molecular marker and DNA sequence information in rice (Oryza sativa L.) has allowed for comparative genetics between rice and wildrice (Zizania palustris L.). In particular, rice simple sequence repeat (SSR) markers were used to develop an SSR-based linkage map in wildrice and to determine the amount of genetic diversity within and among populations of the species. The same set of markers was analyzed in a wildrice population segregating for the pistillate trait to find marker genotypes that correlated with the trait. Rice genome DNA sequences for three well-characterized genes were used to determine the relatedness between Oryza species and wildrice.

Six-hundred-eleven rice SSR markers were tested in wildrice. Approximately 42% of the markers can be amplified in wildrice and about 30% of those markers exhibit polymorphism in the newly developed SRT X BRL mapping population. SRT is a shattering resistant tester line used in the wildrice breeding program. BRL is a breeding line resulting from selection on a population collected from Big Rice Lake in Cass County, Minnesota. An additional 36 SSR markers were developed in Zizania palustris using DNA sequences from the endangered wildrice species Z. texana. About 47% of these markers exhibited co-dominant polymorphism and 23% dominant polymorphism in the SRT X BRL mapping population. In all, seventy-five of these polymorphic markers were used to develop the first SSR-based molecular genetic map of wildrice.

In contrast to the RFLP-based wildrice map, the SSR-based map indicates little synteny between rice and wildrice. This difference may be due to some non-specific amplification with the rice SSR markers. It may also be an indication of the amount of DNA sequence conservation between the two species as the RFLP markers were generated from cDNAs representing expressed DNA sequences. The majority of the rice SSR markers were generated from genomic DNA sequences where SSR motifs are largely in intron or even non-genic sequences. Such regions of eukaryotic genomes are known to be subject to rearrangement which may be reflected in what appears to be a reduced level of detected synteny (Langham et al. 2004).

A molecular genetic marker was identified to be associated with the pistillate gene. Using this genetic marker to select for the pistillate gene would be useful for developing significantly larger mapping populations than are currently possible in wildrice allowing for more precise mapping and QTL studies. In the short-term, use of pistillate wildrice could benefit the wildrice breeding program by increasing cultivar yields as much as 50%. In the long-term, a true breeding pistillate wildrice could facilitate the development of recombinant inbred lines and near isogenic lines for use in producing wildrice hybrids and genetic stocks. Zizania palustris is more closely related to wild Oryza species than to the cultivated species Oryza sativa based on direct comparison of gene sequences from Adhl, Waxy and MatK.

The Rc gene is known to control rice seed pericarp color via the presence of a 14-bp indel sequence. Comparison of the Rc gene sequence among Zizania palustris, Oryza. rufipogon and O. sativa revealed that wildrice contains the indel sequence conferring red pericarp color in O. rufipogon. Further comparative genetics between Z. palustris and wild Oryza species such as O. brachyantha, O. granulata and O. rufipogon may uncover greater synteny than with O. sativa.

Genetic diversity studies of recently collected natural wildrice from lakes in Minnesota and Wisconsin do not appear to correlate with geographic distribution. The assessment of the available genetic diversity in wildrice is important for continued improvements in wildrice breeding. Generally, wildrice genetic diversity analysis has shown that allele uniqueness and frequency are specific to a given natural population. This may be a result of adaptation to the unique environment of each lake. Further diversity analysis using a larger set of populations may lead to a greater understanding of specific alleles involved in wildrice adaptation that could be utilized to produce varieties that would be more uniformly adapted to the varying cultivated conditions in northern Minnesota  .

2009. Identification and characterization of quantitative trait loci for drought resistance in upland rice (Oryza sativa L.).

2009. Identification and characterization of quantitative trait loci for drought resistance in upland rice (Oryza sativa L.).

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ABSTRACT

Breeding improved drought-resistant upland rice cultivars could be accelerated if suitable quantitative trait loci (QTL) with large and consistent effects on grain yield under drought conditions in the desired genetic backgrounds were identified. Experiments were performed with the aim of identifying, characterizing and validating a QTL with the potential to be used in marker-assisted selection. A population from the cross Vandana/Way Rarem was field tested during two consecutive years. A large-effect QTL (qtl!2.1) for grain yield under reproductive-stage drought was identified. This locus had an additive effect equivalent to 47% of the trial mean yield over two years and had a very low level of QTL x year interaction. The favorable effect was contributed by Way Rarem, the drought-susceptible parent. A subset of lines was subsequently tested in a total of 21 trials, in order to confirm the effect of qtl!2.1 under a wider range of environments, stress intensities and stress timing. The QTL had a very consistent effect across trials, as it improved grain yield in 9 out of 10 moderately to severely drought-stressed trials. The locus had no effect under well-watered conditions. The results from those trials confirm that qtl!2.1 is a drought-specific locus that is consistent across environments. Physiological experiments on a subset of lines having contrasting genotypes at qtll2.1 revealed that the Way Rarem-derived allele of qtl!2.1 improves grain yield under drought mainly through a slight improvement (7 to 9 %) in plant water uptake under water-limited conditions. Such an increase in water uptake could explain the large effect on grain yield observed under field conditions. This improvement of plant water uptake is likely associated with improved root architecture. Finally, QTL mapping for drought resistance was also performed in an IR55419-04/ Way Rarem population evaluated under similar conditions as the Vandana/Way Rarem population. This revealed that qtll2.1 had no effect on grain yield under stress in the IR55419-04/Way Rarem population, suggesting QTL x genetic background interaction for qtl!2.1  .


 

2008. Genome-wide analysis of the nitrogen stress transcriptome of red rice (Oryza sativa L.).

2008. Genome-wide analysis of the nitrogen stress transcriptome of red rice (Oryza sativa L.).

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ABTRACT

The weedy red rice (Oryza sativa L.) competes with cultivated rice (O. sativa) for N fertilizers, as demonstrated in field and greenhouse experiments. The ability of red rice to accumulate more N than cultivated rice even at low-N was hypothesized to be due to a molecular mechanism that helps it tolerate N deficiency stress better than the cultivated rice. Microarray analysis of the N stress transcriptome of the weedy red rice was conducted to identify expression patterns that could reveal signaling and response pathways in N deficiency stress adaptation. Candidate genes identified by microarray analysis were validated and their expression patterns in red rice were compared with those in Wells by semi-quantitative real-time RT-PCR. Microarray analysis revealed transcription factors and genes associated with signaling and response regulation, NH4+ assimilation, starch synthesis, stress response, protein fate, and energy metabolism. Gene expression profiles suggested that these genes compose an N deficiency stress response pathway that is likely underlying the greater tolerance of weedy red rice to N-limiting conditions compared to the cultivated rice. Real-time RT-PCR gene assays revealed that genes coding for two transcription factors as well as enzymes involved in starch synthesis, N assimilation, protein folding, and signaling and response regulation were differentially expressed in red rice and cultivated rice. Although these findings confirm the hypothesis that red rice has adaptive molecular mechanisms different from Wells rice, further experiments need to be done to investigate the specific processes involved in the response pathways regulated by these transcription factors and enzymes, particularly at time points immediately after N deficiency  .


 

2008. Genetic and molecular characterization of the low phytic acid (Lpa1) locus in rice

2008. Genetic and molecular characterization of the low phytic acid (Lpa1) locus in rice 

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ABTRACT

The rice low phytic acid (lpa\) mutant has a 45 % reduction in seed phytic acid and a molar equivalent increase in inorganic P. The rice /pal mutation was mapped to rice chromosome 2L. This research aims to identify the rice Lpal gene using a positional cloning and candidate gene approach, characterize the rice Lpal gene expression and identify lpal mutant alleles for functional analysis. High resolution genetic mapping of the rice lpal mutation resulted in the identification of an 8 kb region on chromosome 2 containing a single candidate gene (LOC_Os02g57400) with homology to 2-phosphoglycerate kinase. Sequence analysis of the candidate gene from the two independent mutants (KB 1-1 and DR1331-2) confirmed that (LOC_Os02g57400) is the rice Lpa\ gene {OsLpa 1). This gene has three alternative splice variants which have been confirmed by expression analysis. The location and nature of the two mutations suggests that the lesion affects the translation of the predicted protein derived from the longest transcript. OsLpa 1 represents a novel gene involved in phytic acid metabolism. The identification of Lpa\ gene as well as generation of additional mutants for future study will contribute to the understanding of the phytic acid biosynthesis and accumulation in plants  .