Imaging revealed interesting spaceflight-associated root morphologies. They needed to fix the tissues for further study back on Earth, but conventional fixation methods require separate fixatives depending on whether the sample is intended for molecular or morphological study.
This is a SEM image of Arabidopsis tissue processed using the new single fixation protocol developed by Schultz et al. Pictured is the adaxial leaf epidermis (3500x, scale bar = 8.57 ìm).
Credit: Image courtesy of Schultz et al.
If the scientists wanted to study how spaceflight affected patterns of gene expression central to morphological patterns of cell growth, they needed a fixation method that would allow them to study both perspectives.
Most scientists at work in the laboratory rely on protocols that have been developed without the need for restrictions on the amount of space, time, or reagents they use. For scientists conducting experiments in spaceflight, time and resources are strictly regulated and limited, and researchers must know in advance which protocols will maximize the usefulness of the data they collect.
University of Florida professors Anna-Lisa Paul and Robert Ferl and colleagues are collaborating with the National Aeronautics and Space Administration (NASA) to understand plant growth and development in spaceflight. Along with lead author and graduate student Eric Schultz, they have developed a single fixation protocol for use in space that allows plant material to be used for multiple experimental applications. Their new protocol for sample preparation was tested on Arabidopsis tissues harvested on the International Space Station and is described in the August issue of Applications in Plant Sciences (available for free viewing at http://www.bioone.org/doi/pdf/10.3732/apps.1300034).
Because of limitations in astronaut crew time and orbital resources, previous spaceflight fixation protocols were designated as either molecular or morphological, due to the separate fixatives required for each application. Tissues for morphologic study were fixed in 3% glutaraldehyde (or a similar solution), and tissues for molecular study were fixed in the tissue storage reagent RNAlater. RNAlater has not commonly been used as a morphologic fixative, as it can produce unclear images with high background staining.
The new method developed by Schultz et al. puts RNAlater-fixed samples through a desalination process to return them to a fresh-like state, and then uses low-temperature scanning electron microscopy (cryo-SEM) to preserve tissues for imaging. Because few laboratories have access to the necessary equipment for cryo-SEM, the authors tested and developed a protocol that emulates cryo-SEM using standard SEM equipment and, importantly, that results in minimal tissue damage.
Although it was developed to address specific constraints for spaceflight experiments, Paul notes that their new method is broadly applicable. "There are a lot of situations where biologists want to collect samples in extreme situations. In our case—a space vehicle orbiting the Earth."
The new protocol maximizes the amount of data obtained from a single sample and allows for the concomitant examination of both molecular and morphological features. Using a single fixation protocol, direct comparisons between changes in morphology and altered gene expression can be made. Such an analysis not only makes full use of samples and replicates but also enables a robust analysis of the relationship between heredity and development. "Putting two tools together, it is powerful to look at the morphology in conjunction with the genes that are being expressed," says Paul.
The new protocol boasts low costs and high accessibility, and has wide application to any situation where recovery of biological resources is limited. Notably, this includes researchers collecting and preserving samples in the field, where space for materials is at a premium. "In places where sampling is limited, difficult, or expensive, the use of preservatives allows for more routes to analysis," notes Ferl.
Applications in Plant Sciences (APPS) is a monthly, peer-reviewed, open access journal focusing on new tools, technologies, and protocols in all areas of the plant sciences. It is published by the Botanical Society of America (http://www.botany.org), a non-profit membership society with a mission to promote botany, the field of basic science dealing with the study and inquiry into the form, function, development, diversity, reproduction, evolution, and uses of plants and their interactions within the biosphere. The first issue of APPS published in January 2013; APPS is available as part of BioOne's Open Access collection (http://www.bioone.org/loi/apps).
For further information, please contact the APPS staff at firstname.lastname@example.org.
Beth Parada | EurekAlert!
When Air is in Short Supply - Shedding light on plant stress reactions when oxygen runs short
23.03.2017 | Institut für Pflanzenbiochemie
WPI team grows heart tissue on spinach leaves
23.03.2017 | Worcester Polytechnic Institute
Astronomers from Bonn and Tautenburg in Thuringia (Germany) used the 100-m radio telescope at Effelsberg to observe several galaxy clusters. At the edges of these large accumulations of dark matter, stellar systems (galaxies), hot gas, and charged particles, they found magnetic fields that are exceptionally ordered over distances of many million light years. This makes them the most extended magnetic fields in the universe known so far.
The results will be published on March 22 in the journal „Astronomy & Astrophysics“.
Galaxy clusters are the largest gravitationally bound structures in the universe. With a typical extent of about 10 million light years, i.e. 100 times the...
Researchers at the Goethe University Frankfurt, together with partners from the University of Tübingen in Germany and Queen Mary University as well as Francis Crick Institute from London (UK) have developed a novel technology to decipher the secret ubiquitin code.
Ubiquitin is a small protein that can be linked to other cellular proteins, thereby controlling and modulating their functions. The attachment occurs in many...
In the eternal search for next generation high-efficiency solar cells and LEDs, scientists at Los Alamos National Laboratory and their partners are creating...
Silicon nanosheets are thin, two-dimensional layers with exceptional optoelectronic properties very similar to those of graphene. Albeit, the nanosheets are less stable. Now researchers at the Technical University of Munich (TUM) have, for the first time ever, produced a composite material combining silicon nanosheets and a polymer that is both UV-resistant and easy to process. This brings the scientists a significant step closer to industrial applications like flexible displays and photosensors.
Silicon nanosheets are thin, two-dimensional layers with exceptional optoelectronic properties very similar to those of graphene. Albeit, the nanosheets are...
Enzymes behave differently in a test tube compared with the molecular scrum of a living cell. Chemists from the University of Basel have now been able to simulate these confined natural conditions in artificial vesicles for the first time. As reported in the academic journal Small, the results are offering better insight into the development of nanoreactors and artificial organelles.
Enzymes behave differently in a test tube compared with the molecular scrum of a living cell. Chemists from the University of Basel have now been able to...
20.03.2017 | Event News
14.03.2017 | Event News
07.03.2017 | Event News
23.03.2017 | Life Sciences
23.03.2017 | Power and Electrical Engineering
23.03.2017 | Earth Sciences