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Plant Methods (1)
Edelmann, Richard E. (1)
Johnson, Christina M. (1)
Kiss, John Z (1)
Kiss, John Z. (1)
Millar, Katherine D.L. (1)
Molas, M Lia (1)
Year of Publication
An Endogenous Growth Pattern of Roots Is Revealed in Seedlings Grown in Microgravity
Millar, Katherine D.L.
Johnson, Christina M.
Edelmann, Richard E.
In plants, sensitive and selective mechanisms have evolved to perceive and respond to light and gravity. We investigated the effects of microgravity on the growth and development of Arabidopsis thaliana (ecotype Landsberg) in a spaceflight experiment. These studies were performed with the Biological Research in Canisters (BRIC) hardware system in the middeck region of the space shuttle during mission STS-131 in April 2010. Seedlings were grown on nutrient agar in Petri dishes in BRIC hardware under dark conditions and then fixed in flight with paraformaldehyde, glutaraldehyde, or RNAlater. Although the long-term objective was to study the role of the actin cytoskeleton in gravity perception, in this article we focus on the analysis of morphology of seedlings that developed in microgravity. While previous spaceflight studies noted deleterious morphological effects due to the accumulation of ethylene gas, no such effects were observed in seedlings grown with the BRIC system. Seed germination was 89% in the spaceflight experiment and 91% in the ground control, and seedlings grew equally well in both conditions. However, roots of space-grown seedlings exhibited a significant difference (compared to the ground controls) in overall growth patterns in that they skewed to one direction. In addition, a greater number of adventitious roots formed from the axis of the hypocotyls in the flight-grown plants. Our hypothesis is that an endogenous response in plants causes the roots to skew and that this default growth response is largely masked by the normal 1 g conditions on Earth. Key Words: Gravity—Multicellular life—Spacecraft experiments—Spaceflight. Astrobiology 11, 787–797.
The effect of column purification on cDNA indirect labelling for microarrays
Molas, M Lia
The success of the microarray reproducibility is dependent upon the performance of standardized procedures. Since the introduction of microarray technology for the analysis of global gene expression, reproducibility of results among different laboratories has been a major problem. Two of the main contributors to this variability are the use of different microarray platforms and different laboratory practices. In this paper, we address the latter question in terms of how variation in one of the steps of a labelling procedure affects the cDNA product prior to microarray hybridization.
We used a standard procedure to label cDNA for microarray hybridization and employed different types of column chromatography for cDNA purification. After purifying labelled cDNA, we used the Agilent 2100 Bioanalyzer and agarose gel electrophoresis to assess the quality of the labelled cDNA before its hybridization onto a microarray platform. There were major differences in the cDNA profile (i.e. cDNA fragment lengths and abundance) as a result of using four different columns for purification. In addition, different columns have different efficiencies to remove rRNA contamination. This study indicates that the appropriate column to use in this type of protocol has to be experimentally determined. Finally, we present new evidence establishing the importance of testing the method of purification used during an indirect labelling procedure. Our results confirm the importance of assessing the quality of the sample in the labelling procedure prior to hybridization onto a microarray platform.
Standardization of column purification systems to be used in labelling procedures will improve the reproducibility of microarray results among different laboratories. In addition, implementation of a quality control check point of the labelled samples prior to microarray hybridization will prevent hybridizing a poor quality sample to expensive micorarrays.
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