HydroLink gel electrophoresis (HLGE). III. High DNA loading capacity and recovery of dsDNA.

Novel polymers have been prepared for high performance electrophoretic separations of double-stranded DNA (dsDNA). These materials are part of a family of HydroLink high performance electrophoresis polymers. A comparison of the resolving capabilities of dsDNA HydroLink gels to agarose and polyacrylamide separations has been described in an accompanying paper. In this study, we demonstrate that dsDNA HydroLink gels possess ten times the loading capacity of comparable polyacrylamide or agarose gels without compromise to resolution or biological integrity of the separated DNA. A simplified procedure for recovery of separated components is also described.

HydroLink gel electrophoresis (HLGE). II. Applications of a new polymer matrix to dsDNA analysis.

HydroLink materials represent a novel family of gels composed of unique polymer matrices. The applications of HydroLink to molecular biology and, specifically, to DNA technology have been carefully investigated. Our results indicate that the HydroLink matrix developed for double-stranded DNA (dsDNA) is an excellent tool for electrophoretic separations in fixed electric fields. Excellent linear resolution from 100 to 5000 base pairs is easily achieved with good resolution albeit non-linear from 6000 to 23000 base pairs. The broad range of separation in addition to increased mechanical strength of dsDNA HydroLink represents a distinct advantage over other matrices currently used in DNA electrophoretic analysis.

Science potential from a Europa lander.

The prospect of a future soft landing on the surface of Europa is enticing, as it would create science opportunities that could not be achieved through flyby or orbital remote sensing, with direct relevance to Europa's potential habitability. Here, we summarize the science of a Europa lander concept, as developed by our NASA-commissioned Science Definition Team. The science concept concentrates on observations that can best be achieved by in situ examination of Europa from its surface. We discuss the suggested science objectives and investigations for a Europa lander mission, along with a model planning payload of instruments that could address these objectives. The highest priority is active sampling of Europa's non-ice material from at least two different depths (0.5-2 cm and 5-10 cm) to understand its detailed composition and chemistry and the specific nature of salts, any organic materials, and other contaminants. A secondary focus is geophysical prospecting of Europa, through seismology and magnetometry, to probe the satellite's ice shell and ocean. Finally, the surface geology can be characterized in situ at a human scale. A Europa lander could take advantage of the complex radiation environment of the satellite, landing where modeling suggests that radiation is about an order of magnitude less intense than in other regions. However, to choose a landing site that is safe and would yield the maximum science return, thorough reconnaissance of Europa would be required prior to selecting a scientifically optimized landing site.

H2O at the Phoenix landing site.

The Phoenix mission investigated patterned ground and weather in the northern arctic region of Mars for 5 months starting 25 May 2008 (solar longitude between 76.5 degrees and 148 degrees ). A shallow ice table was uncovered by the robotic arm in the center and edge of a nearby polygon at depths of 5 to 18 centimeters. In late summer, snowfall and frost blanketed the surface at night; H(2)O ice and vapor constantly interacted with the soil. The soil was alkaline (pH = 7.7) and contained CaCO(3), aqueous minerals, and salts up to several weight percent in the indurated surface soil. Their formation likely required the presence of water.

A closer look at water-related geologic activity on Mars.

Water has supposedly marked the surface of Mars and produced characteristic landforms. To understand the history of water on Mars, we take a close look at key locations with the High-Resolution Imaging Science Experiment on board the Mars Reconnaissance Orbiter, reaching fine spatial scales of 25 to 32 centimeters per pixel. Boulders ranging up to approximately 2 meters in diameter are ubiquitous in the middle to high latitudes, which include deposits previously interpreted as finegrained ocean sediments or dusty snow. Bright gully deposits identify six locations with very recent activity, but these lie on steep (20 degrees to 35 degrees) slopes where dry mass wasting could occur. Thus, we cannot confirm the reality of ancient oceans or water in active gullies but do see evidence of fluvial modification of geologically recent mid-latitude gullies and equatorial impact craters.

Soil temperatures and stability of ice-cemented ground in the McMurdo Dry Valleys, Antarctica.

Year-round temperature measurements at 1600 m elevation during 1994 in the Asgard Range Antarctica, indicate that the mean annual frost point of the ice-cemented ground, 25 cm below the surface, is -21.7 +/- 0.2 degrees C and the mean annual frost point of the atmosphere is -27.5 +/- 1.0 degrees C. The corresponding mean annual temperatures are -24.9 degrees C and -23.3 degrees C. These results imply that there is a net flux of water vapour from the ice to the atmosphere resulting in a recession of the ice-cemented ground by about 0.4-0.6 mm yr-1. The level of the ice-cemented permafrost is about 12 cm below the level of dry permafrost. The summer air temperatures would have to increase about 7 degrees C for thawing temperatures to just reach the top of the subsurface ice. Either subsurface ice at this location is evaporating over time or there are sporadic processes that recharge the ice and maintain equilibrium over long timescales.

New nitrogen-fixing microorganisms detected in oligotrophic oceans by amplification of Nitrogenase (nifH) genes.

Oligotrophic oceanic waters of the central ocean gyres typically have extremely low dissolved fixed inorganic nitrogen concentrations, but few nitrogen-fixing microorganisms from the oceanic environment have been cultivated. Nitrogenase gene (nifH) sequences amplified directly from oceanic waters showed that the open ocean contains more diverse diazotrophic microbial populations and more diverse habitats for nitrogen fixers than previously observed by classical microbiological techniques. Nitrogenase genes derived from unicellular and filamentous cyanobacteria, as well as from the alpha and gamma subdivisions of the class Proteobacteria, were found in both the Atlantic and Pacific oceans. nifH sequences that cluster phylogenetically with sequences from sulfate reducers or clostridia were found associated with planktonic crustaceans. Nitrogenase sequence types obtained from invertebrates represented phylotypes distinct from the phylotypes detected in the picoplankton size fraction. The results indicate that there are in the oceanic environment several distinct potentially nitrogen-fixing microbial assemblages that include representatives of diverse phylotypes.

New Nitrogen-Fixing Microorganisms Detected in Oligotrophic Oceans by Amplification of Nitrogenase (nifH) Genes.

[This corrects the article on p. 3444 in vol. 64, PMID: 9726895.].

Expression of nifH genes in natural microbial assemblages in Lake George, New York, detected by reverse transcriptase PCR.

A modified nested reverse transcriptase PCR (RT-PCR) method was used to detect the expression of nitrogenase genes in meso-oligotrophic Lake George, New York. Net (>20-microm pore size) plankton samples collected from two sites (Dome Island and Hague Marina) were extracted for total RNA and genomic DNA to determine the identity of diazotrophic organisms that were present and those that were actively expressing nitrogenase genes. Phylogenetic analysis of individual sequences cloned from PCR amplifications showed that there were phylogenetically diverse groups of bacteria that possessed a nifH gene, including representatives of unicellular and filamentous cyanobacteria, the alpha- and gamma-subdivisions of the division Proteobacteria (alpha- and gamma-proteobacteria), and a previously undefined group of bacteria. The phylotypes cloned from RT-PCR amplifications, which were actively expressing nifH transcripts, clustered with the unicellular and filamentous cyanobacteria, alpha-proteobacteria, and the novel bacterial cluster. No bacterial sequences were found which clustered with sequences from cluster II (alternative nitrogenases), III (nitrogenases in strict anaerobes), or IV (nifH-like sequences). These results indicate that there were several distinct groups of nitrogen-fixing microorganisms in the net plankton from both sampling sites and that most of the groups had representative phylotypes that were actively expressing nitrogenase genes.

Circadian rhythm of nitrogenase gene expression in the diazotrophic filamentous nonheterocystous cyanobacterium Trichodesmium sp. strain IMS 101.

Recent studies suggested that the daily cycle of nitrogen fixation activity in the marine filamentous nonheterocystous cyanobacterium Trichodesmium sp. is controlled by a circadian rhythm. In this study, we evaluated the rhythm of nitrogen fixation in Trichodesmium sp. strain IMS 101 by using the three criteria for an endogenous rhythm. Nitrogenase transcript abundance oscillated with a period of approximately 24 h, and the cycle was maintained even under constant light conditions. The cyclic pattern of transcript abundance was maintained when the culture was grown at 24 and 28.5 degrees C, although the period was slightly longer (26 h) at the higher temperature. The cycle of gene expression could be entrained with light-dark cues. Results of inhibitor experiments indicated that transcript abundance was regulated primarily by transcription initiation, rather than by degradation. The circadian rhythm, the first conclusively demonstrated endogenous rhythm in a filamentous cyanobacterium, was also reflected in nitrogenase MoFe protein abundance and patterns of Fe protein posttranslational modification-demodification.

Expression of photosynthesis genes in relation to nitrogen fixation in the diazotrophic filamentous nonheterocystous cyanobacterium Trichodesmium sp. IMS 101.

The daily cycle of nitrogenase expression in the marine filamentous nonheterocystous cyanobacterium Trichodesmium spp. is controlled by a circadian rhythm. We evaluated the rhythm of two key photosynthesis genes, psbA of photosystem II and psaA of photosystem I, in Trichodesmium sp. IMS 101 using the 3 criteria for an endogenous rhythm. The transcript abundance of psbA and psaA transcripts oscillated with a period of ca. 24 h under a 12 h light/12 h dark regime. At 24 degrees C and 28 degrees C the cyclic pattern of transcript abundance was maintained for at least 58 h under constant light conditions, whereas the periods were about 24 h at 24 degrees C, and 26-30 h at the higher temperature. The cycles of psbA and psaA gene expression were entrained using light-dark cues. Transcription of nifHDK was initiated prior to the light period, followed by psbA and finally psaA. There was a 90 degrees (6 h) phase difference between the net accumulation of nifHDK and psbA transcripts, as well as between that of psbA and psaA transcripts. Results of inhibitor experiments indicated that psbA and psaA transcription was regulated differently by initiation and degradation during the light period. Short-term changes of light conditions resulted in significant effects on psbA transcription and nitrogenase activity, but had less of an effect on psaA and nifHDK transcription.