Analysis of anther transcriptomes to identify genes contributing to meiosis and male gametophyte development in rice

Extent: 20p.Background: In flowering plants, the anther is the site of male gametophyte development. Two major events in the development of the male germline are meiosis and the asymmetric division in the male gametophyte that gives rise to the vegetative and generative cells, and the following mitotic division in the generative cell that produces two sperm cells. Anther transcriptomes have been analyzed in many plant species at progressive stages of development by using microarray and sequence-by synthesis-technologies to identify genes that regulate anther development. Here we report a comprehensive analysis of rice anther transcriptomes at four distinct stages, focusing on identifying regulatory components that contribute to male meiosis and germline development. Further, these transcriptomes have been compared with the transcriptomes of 10 stages of rice vegetative and seed development to identify genes that express specifically during anther development. Results: Transcriptome profiling of four stages of anther development in rice including pre-meiotic (PMA), meiotic (MA), anthers at single-celled (SCP) and tri-nucleate pollen (TPA) revealed about 22,000 genes expressing in at least one of the anther developmental stages, with the highest number in MA (18,090) and the lowest (15,465) in TPA. Comparison of these transcriptome profiles to an in-house generated microarray-based transcriptomics database comprising of 10 stages/tissues of vegetative as well as reproductive development in rice resulted in the identification of 1,000 genes specifically expressed in anther stages. From this sub-set, 453 genes were specific to TPA, while 78 and 184 genes were expressed specifically in MA and SCP, respectively. The expression pattern of selected genes has been validated using real time PCR and in situ hybridizations. Gene ontology and pathway analysis of stage-specific genes revealed that those encoding transcription factors and components of protein folding, sorting and degradation pathway genes dominated in MA, whereas in TPA, those coding for cell structure and signal transduction components were in abundance. Interestingly, about 50% of the genes with anther-specific expression have not been annotated so far. Conclusions: Not only have we provided the transcriptome constituents of four landmark stages of anther development in rice but we have also identified genes that express exclusively in these stages. It is likely that many of these candidates may therefore contribute to specific aspects of anther and/or male gametophyte development in rice. In addition, the gene sets that have been produced will assist the plant reproductive community in building a deeper understanding of underlying regulatory networks and in selecting gene candidates for functional validation.Priyanka Deveshwar, William D Bovill, Rita Sharma, Jason A Able and Sanjay Kapoo

Methodology for High-Throughput Field Phenotyping of Canopy Temperature Using Airborne Thermography

Lower canopy temperature (CT), resulting from increased stomatal conductance, has been associated with increased yield in wheat. Historically, CT has been measured with hand-held infrared thermometers. Using the hand-held CT method on large field trials is problematic, mostly because measurements are confounded by temporal weather changes during the time required to measure all plots. The hand-held CT method is laborious and yet the resulting heritability low, thereby reducing confidence in selection in large scale breeding endeavors. We have developed a reliable and scalable crop phenotyping method for assessing CT in large field experiments. The method involves airborne thermography from a manned helicopter using a radiometrically-calibrated thermal camera. Thermal image data is acquired from large experiments in the order of seconds, thereby enabling simultaneous measurement of CT on potentially 1000s of plots. Effects of temporal weather variation when phenotyping large experiments using hand-held infrared thermometers are therefore reduced. The method is designed for cost-effective and large-scale use by the non-technical user and includes custom-developed software for data processing to obtain CT data on a single-plot basis for analysis. Broad-sense heritability was routinely >0.50, and as high as 0.79, for airborne thermography CT measured near anthesis on a wheat experiment comprising 768 plots of size 2 × 6 m. Image analysis based on the frequency distribution of temperature pixels to remove the possible influence of background soil did not improve broad-sense heritability. Total image acquisition and processing time was ca. 25 min and required only one person (excluding the helicopter pilot). The results indicate the potential to phenotype CT on large populations in genetics studies or for selection within a plant breeding program.This research was funded by the Australian Government National Collaborative Research Infrastructure Strategy (Australian Plant Phenomics Facility) and the Grains Research and Development Corporation (GRDC)

An autoactive NB-LRR gene causes Rht13 dwarfism in wheat

Semidwarfing genes have greatly increased wheat yields globally, yet the widely used gibberellin (GA)-insensitive genes Rht-B1b and Rht-D1b have disadvantages for seedling emergence. Use of the GA-sensitive semidwarfing gene Rht13 avoids this pleiotropic effect. Here, we show that Rht13 encodes a nucleotide-binding site/leucine-rich repeat (NB-LRR) gene. A point mutation in the semidwarf Rht-B13b allele autoactivates the NB-LRR gene and causes a height reduction comparable with Rht-B1b and Rht-D1b in diverse genetic backgrounds. The autoactive Rht-B13b allele leads to transcriptional up-regulation of pathogenesis-related genes including class III peroxidases associated with cell wall remodeling. Rht13 represents a new class of reduced height (Rht) gene, unlike other Rht genes, which encode components of the GA signaling or metabolic pathways. This discovery opens avenues to use autoactive NB-LRR genes as semidwarfing genes in a range of crop species, and to apply Rht13 in wheat breeding programs using a perfect genetic marker

Identification, validation, and pyramiding of quantitative trait loci for resistance to crown rot in wheat

[Abstract]: Crown rot (causal organism: Fusarium pseudograminearum) is a significant disease affecting wheat in Australia. Although first reported over 60 years ago, the disease has become more prevalent in recent years due to the adoption of minimum tillage and stubble retention practices. Breeding for resistance to crown rot is difficult - phenotypic selection, which is usually done at harvest, is time-consuming, expensive, and subject to between year variability due to sensitivity to environmental conditions. For these reasons, the coupling of molecular techniques with conventional plant breeding (marker-assisted selection) has the potential to more rapidly and reliably identify genomic regions that contribute to resistance. The objective of this study was to identify, validate, and pyramid quantitative trait loci (QTL) for resistance to crown rot present in a W21MMT70 x Mendos doubled haploid wheat population. Replicated seedling trials were conducted in 2001, 2003, and 2005. In each seedling trial, W21MMT70 displayed partial resistance to crown rot whereas Mendos seedlings were susceptible. A bulked segregant analysis (BSA), using 390 simple sequence repeat (SSR) markers chosen for their coverage of the wheat genome, was initially conducted based upon the 2001 seedling trial data in an attempt to rapidly identify genomic regions associated to resistance. The BSA did not reveal any markers associated with resistance to crown rot. As a result, a full mapping study was conducted. One hundred and twenty eight (128) SSR markers were mapped across the population to produce a framework map. Previously screened AFLP markers were added to the map. Composite interval mapping revealed eight QTL associated with resistance. Of these, three (located on chromosomes 2B, 2D, and 5D) were consistently detected in each of the three seedling trials. Two QTL (on chromosomes 1A and 3B) were detected in two of the three trials. The 2D, 3B, and 5D QTL were inherited from W21MMT70, whereas the 1A and 2B QTL were inherited from Mendos. Two software programs were used to identify epistatic interactions between QTL. While the results of the two programs differed markedly, both programs detected a highly significant interaction between the W21MMT70 inherited 5D QTL and a locus on chromosome 2D inherited from Mendos. The overall effect of the epistatic interactions was not as great as the additive effects of nonepistatic QTL. Nonetheless, the presence of epistasis may indicate that, particularly in the case of 5D, the effect of this QTL may be dependent on the background into which it is introgressed. Validation of three W21MMT70-inherited QTL (on chromosomes 2D, 3B, and 5D) was conducted on three F2 populations with W21MMT70 as one of the parents. While the 5D QTL was validated in two of the three crosses, neither the 2D nor the 3B QTL were detected in any of the F2 validation populations. It is likely that the size of the F2 populations (the largest composed of 94 individuals), in conjunction with the variability that is inherent when screening for resistance to crown rot, precluded validation of these regions. Validation of the 2B Mendos-inherited QTL was conducted on a Sunco x Batavia doubled haploid population because Sunco possesses the same Triticum timopheevi 2B introgression that is present in Mendos. This validated QTL (designated Q.CR..usq-2B2) explained 11 % of the phenotypic variance in the Sunco x Batavia population. To assess the effectiveness of pyramiding QTL for resistance to crown rot, a 2-49 x W21MMT70 population was examined. A number of lines of this population performed significantly better than each of the parents in the replicated seedling trial that was conducted. Four QTL, located on chromosomes 1A, 1D, 2D, and 3B, were detected. The 1A and 1D QTL were inherited from 2-49 whereas the 2D and 3B QTL were inherited from W21MMT70. The 1A QTL from 2-49 has not been previously validated, and this QTL has been designated QCr.usq-1A1. The 3B QTL (designated QCr.usq-3B1) had the highest effect (LRS 42.1; explaining 21.0 % of the phenotypic variance) in the 2-49 x W21MMT70 population. The 2D QTL (QCr.usq-2D1) was shown to have a minor effect. The 5D QTL that was inherited from W21MMT70 in the W21MMT70 x Mendos population was not detected in the 2-49 x W21MMT70 population. A number of possible explanations for the inability to detect this QTL in the 2-49 x W21MMT70 population are discussed

Guide To Environmental & Spatial Variables

Description of environmental and spatial variables including units of measuremen

Environmental & Spatial Variables

Various measures of environmental variation at 8 sites in each of two streams during summer plus the latitude and longitude for each site

Data from: A fresh approach reveals how dispersal shapes metacommunity structure in a human-altered landscape

1. To understand species losses from disturbed landscapes, it is important to distinguish the effects of degraded environmental conditions from those caused by barriers to dispersal between habitat patches. To assess the relative importance of these effects, we developed a new approach using permutation and association tests applied to rank abundance data, using the invertebrate fauna of two rivers in two seasons. 2. Our study streams were Hughes Creek and Seven Creeks, in south-eastern Australia, which have both been degraded by agriculture in downstream sections. We collected benthic invertebrates and also dispersing individuals (drift, terrestrial adults) during two seasons in 2007–2008. Study sites spanned strong environmental gradients as well as the main dispersal route (up- and down-channel). Environmental data were analysed to set up permutation tests on rank abundances. Survey and disperser data were contrasted using contingency table analyses. 3. The results suggest dispersal plays a strong role in community structure. Environmental effects were evident and strongest upstream, but evidence of environmental effects was weak over much of the gradient. Many species had different distributions in different data sets or dispersers that were abundant at locations distant from centres of benthic distribution. 4. Our results differ from many studies, but few have been able to evaluate dispersal effects directly. Our method provides a practical approach for evaluating the role dispersal plays in driving species abundance patterns across landscapes, thus bridging a gap between theory and practice. 5. Synthesis and applications. Managers typically use indices of ecosystem health that assume environmental conditions largely determine species diversity and abundance. Dispersal between habitat patches is known to be important, but there are no reliable methods to assess the role dispersal may play. We provide an approach that allows both dispersal and environmental effects on species distributions to be evaluated from survey data. This may open the way for dispersal information to be incorporated into management actions. Additionally, the approach should allow improved siting of restoration projects that depend greatly on successful dispersal of individuals for successful outcomes

Drift Data

Average numbers or individuals in the drift (over two hours) of multiple species of invertebrates sampled at 4 sites in each of two streams and two seasons

List Of Species Names

A list of all species in the benthic and drift data sets with taxonomic classification and variable names

Water & Air Temperatures

Air and water temperatures recorded at two sites on each of Hughes Creek and Seven Creeks during summer and spring in 2008 and 2009