Genome-wide transcriptome analysis of the transition from primary to secondary stem development in Populus trichocarpa

<p>Abstract</p> <p>Background</p> <p>With its genome sequence and other experimental attributes, <it>Populus trichocarpa </it>has become the model species for genomic studies of wood development. Wood is derived from secondary growth of tree stems, and begins with the development of a ring of vascular cambium in the young developing stem. The terminal region of the developing shoot provides a steep developmental gradient from primary to secondary growth that facilitates identification of genes that play specialized functions during each of these phases of growth.</p> <p>Results</p> <p>Using a genomic microarray representing the majority of the transcriptome, we profiled gene expression in stem segments that spanned primary to secondary growth. We found 3,016 genes that were differentially expressed during stem development (Q-value ≤ 0.05; >2-fold expression variation), and 15% of these genes encode proteins with no significant identities to known genes. We identified all gene family members putatively involved in secondary growth for carbohydrate active enzymes, tubulins, actins, actin depolymerizing factors, fasciclin-like AGPs, and vascular development-associated transcription factors. Almost 70% of expressed transcription factors were upregulated during the transition to secondary growth. The primary shoot elongation region of the stem contained specific carbohydrate active enzyme and expansin family members that are likely to function in primary cell wall synthesis and modification. Genes involved in plant defense and protective functions were also dominant in the primary growth region.</p> <p>Conclusion</p> <p>Our results describe the global patterns of gene expression that occur during the transition from primary to secondary stem growth. We were able to identify three major patterns of gene expression and over-represented gene ontology categories during stem development. The new regulatory factors and cell wall biogenesis genes that we identified provide candidate genes for further functional characterization, as well as new tools for molecular breeding and biotechnology aimed at improvement of tree growth rate, crown form, and wood quality.</p

Validating internal controls for quantitative plant gene expression studies

BACKGROUND: Real-time reverse transcription PCR (RT-PCR) has greatly improved the ease and sensitivity of quantitative gene expression studies. However, accurate measurement of gene expression with this method relies on the choice of a valid reference for data normalization. Studies rarely verify that gene expression levels for reference genes are adequately consistent among the samples used, nor compare alternative genes to assess which are most reliable for the experimental conditions analyzed. RESULTS: Using real-time RT-PCR to study the expression of 10 poplar (genus Populus) housekeeping genes, we demonstrate a simple method for determining the degree of stability of gene expression over a set of experimental conditions. Based on a traditional method for analyzing the stability of varieties in plant breeding, it defines measures of gene expression stability from analysis of variance (ANOVA) and linear regression. We found that the potential internal control genes differed widely in their expression stability over the different tissues, developmental stages and environmental conditions studied. CONCLUSION: Our results support that quantitative comparisons of candidate reference genes are an important part of real-time RT-PCR studies that seek to precisely evaluate variation in gene expression. The method we demonstrated facilitates statistical and graphical evaluation of gene expression stability. Selection of the best reference gene for a given set of experimental conditions should enable detection of biologically significant changes in gene expression that are too small to be revealed by less precise methods, or when highly variable reference genes are unknowingly used in real-time RT-PCR experiments

Structure and expression of two Populus trichocarpa homologs of the floral homeotic gene AGAMOUS

Because of their small genomes, facile clonal propagation, fast growth, and susceptibility to Agrobacterium transformation, poplars (genus Populus) are widely considered model systems for the application of molecular genetics and biotechnology in forestry. However, a major concern over commercial use of genetically engineered trees is the release of transgenes into wild populations. The goal of this study was to characterize two genes that are expected to be critical for female and male reproductive development in poplars, and thus, could be used for genetic engineering of reproductive sterility. This trait would mitigate ecological risks associated with commercial deployment of transgenic trees by preventing the spread of transgenes via pollen and seed. A secondary goal was to use these genes as probes to help understand the reproductive biology of poplars, whose two-whorled, unisexual flowers are distinct from those of any previously studied plant species. We isolated and characterized two closely related genes from P. trichocarpa (black cottonwood), a native tree of the Pacific Northwest. These genes are homologous to AGAMOUS (AG), a gene controlling reproductive development in the model herbaceous plant Arabidopsis. The proteins encoded by PTAG1 and PTAG2 are 89% identical, and phylogenetic analysis suggests that they are most closely related to genes which specify the identity of both stamens and carpels in herbaceous plants. Gene structure is conserved between PTAG1, PTAG2, AG, and the Antirrhinum AG ortholog, PLENA. The floral RNA expression patterns of the PTAG genes are also very similar to those of AG and PLENA. In situ hybridization studies revealed that PTAG1 and PTAG2 are expressed in the center of both female and male floral meristems before reproductive organ primordia have initiated, and in developing stamen and carpels. Unlike AG, PLENA, and other close AG homologs, PTAG transcripts are detected in vegetative tissue. These results suggest that PTAG1 and PTAG2 may function in a largely redundant manner to specify reproductive organs in Populus. Therefore, inhibiting the endogenous genes or proteins is likely to be an effective way to genetically engineer reproductive sterility. However, weak vegetative expression may preclude use of their promoters to ablate floral tissues

Factors Affecting Implementation of the California Childhood Obesity Research Demonstration (CA-CORD) Project, 2013.

IntroductionEcological approaches to health behavior change require effective engagement from and coordination of activities among diverse community stakeholders. We identified facilitators of and barriers to implementation experienced by project leaders and key stakeholders involved in the Imperial County, California, Childhood Obesity Research Demonstration project, a multilevel, multisector intervention to prevent and control childhood obesity.MethodsA total of 74 semistructured interviews were conducted with project leaders (n = 6) and key stakeholders (n = 68) representing multiple levels of influence in the health care, early care and education, and school sectors. Interviews, informed by the Multilevel Implementation Framework, were conducted in 2013, approximately 12 months after year-one project implementation, and were transcribed, coded, and summarized.ResultsRespondents emphasized the importance of engaging parents and of ensuring support from senior leaders of participating organizations. In schools, obtaining teacher buy-in was described as particularly important, given lower perceived compatibility of the intervention with organizational priorities. From a program planning perspective, key facilitators of implementation in all 3 sectors included taking a participatory approach to the development of program materials, gradually introducing intervention activities, and minimizing staff burden. Barriers to implementation were staff turnover, limited local control over food provided by external vendors or school district policies, and limited availability of supportive resources within the broader community.ConclusionProject leaders and stakeholders in all sectors reported similar facilitators of and barriers to implementation, suggesting the possibility for synergy in intervention planning efforts

Genome-wide analysis of Aux/IAA and ARF gene families in Populus trichocarpa

<p>Abstract</p> <p>Background</p> <p>Auxin/Indole-3-Acetic Acid (Aux/IAA) and Auxin Response Factor (ARF) transcription factors are key regulators of auxin responses in plants. We identified the suites of genes in the two gene families in <it>Populus </it>and performed comparative genomic analysis with <it>Arabidopsis </it>and rice.</p> <p>Results</p> <p>A total of 35 <it>Aux/IAA </it>and 39 <it>ARF </it>genes were identified in the <it>Populus </it>genome. Comparative phylogenetic analysis revealed that several Aux/IAA and ARF subgroups have differentially expanded or contracted between the two dicotyledonous plants. Activator <it>ARF </it>genes were found to be two fold-overrepresented in the <it>Populus </it>genome. <it>PoptrIAA </it>and <it>PoptrARF </it>gene families appear to have expanded due to high segmental and low tandem duplication events. Furthermore, expression studies showed that genes in the expanded <it>PoptrIAA3 </it>subgroup display differential expression.</p> <p>Conclusion</p> <p>The present study examines the extent of conservation and divergence in the structure and evolution of <it>Populus Aux/IAA </it>and <it>ARF </it>gene families with respect to <it>Arabidopsis </it>and rice. The gene-family analysis reported here will be useful in conducting future functional genomics studies to understand how the molecular roles of these large gene families translate into a diversity of biologically meaningful auxin effects.</p

Dependent infrastructure system modeling: A case study of the St. Kitts power and water distribution systems

Critical infrastructure systems underlie the economy, national security, and health of modern society. These infrastructures have become increasingly dependent on each other, which poses challenges when modeling these systems. Although a number of methods have been developed for this problem, few case studies that model real-world dependent infrastructures have been conducted. In this paper, we aim to provide another example of such a case study by modeling a real-world water distribution system dependent on a power system. Unlike in the limited previous case studies, our case study is in a developing nation context. This makes the availability of data about the infrastructure systems in this case study very limited, which is a common characteristic of real-world studies in many settings. Thus, a main contribution of the paper is to show how one can still develop representative, useful models for systems in the context of limited data. To demonstrate the utility of these types of models, two examples of different analyses are performed, where the results provide information about the most vulnerable parts of the infrastructures and critical linkages between the power and water distribution systems.publishedVersio

Evolved Gas Measurements Planned for the Lower Layers of the Gale Crater Mound with the Sample Analysis at Mars Instrument Suite

The lower mound strata of Gale Crater provide a diverse set of chemical environments for exploration by the varied tools of the Curiosity Rover of the Mars Science Laboratory (MSL) Mission. Orbital imaging and spectroscopy clearly reveal distinct layers of hydrated minerals, sulfates, and clays with abundant evidence of a variety of fluvial processes. The three instruments of the MSL Sample Analysis at aMars (SAM) investigation, the Quadrupole Mass Spectrometer (QMS), the Tunable Laser Spectrometer (TLS), and the Gas Chromatograph (GC) are designed to analyze either atmospheric gases or volatiles thermally evolved or chemically extracted from powdered rock or soil. The presence or absence of organic compounds in these layers is of great interest since such an in situ search for this type of record has not been successfully implemented since the mid-60s Viking GCMS experiments. However, regardless of the outcome of the analysis for organics, the abundance and isotopic composition of thermally evolved inorganic compounds should also provide a rich data set to complement the mineralogical and elemental information provided by other MSL instruments. In addition, these evolved gas analysis (EGA) experiments will help test sedimentary models proposed by Malin and Edgett (2000) and then further developed by Milliken et al (2010) for Gale Crater. In the SAM EGA experiments the evolution temperatures of H2O, CO2, SO2, O2, or other simple compounds as the samples are heated in a helium stream to 1000 C provides information on mineral types and their associations. The isotopic composition of O, H, C, and S can be precisely determined in several evolved compounds and compared with the present day atmosphere. Such SAM results might be able to test mineralogical evidence of changing sedimentary and alteration processes over an extended period of time. For example, Bibring et al (2006) have suggested such a major shift from early nonacidic to later acidic alteration. We will illustrate through a variety of evolved gas experiments implemented under SAM-like gas flow and temperature ramp conditions on terrestrial analog minerals on high fidelity Sam breadboards the type of chemical information we expect SAM to provide

Strategies for Engineering Reproductive Sterility in Plantation Forests

A considerable body of research exists concerning the development of technologies to engineer sterility in forest trees. The primary driver for this work has been to mitigate concerns arising from gene flow from commercial plantings of genetically engineered (GE) trees to non-GE plantations, or to wild or feral relatives. More recently, there has been interest in the use of sterility technologies as a means to mitigate the global environmental and socio-economic damage caused by the escape of non-native invasive tree species from planted forests. The current sophisticated understanding of the molecular processes underpinning sexual reproduction in angiosperms has facilitated the successful demonstration of a number of control strategies in hardwood tree species, particularly in the model hardwood tree Poplar. Despite gymnosperm softwood trees, such as pines, making up the majority of the global planted forest estate, only pollen sterility, via cell ablation, has been demonstrated in softwoods. Progress has been limited by the lack of an endogenous model system, long timescales required for testing, and key differences between softwood reproductive pathways and those of well characterized angiosperm model systems. The availability of comprehensive genome and transcriptome resources has allowed unprecedented insights into the reproductive processes of both hardwood and softwood tree species. This increased fundamental knowledge together with the implementation of new breeding technologies, such as gene editing, which potentially face a less oppressive regulatory regime, is making the implementation of engineered sterility into commercial forestry a realistic possibility