Multi-node graphs: a framework for multiplexed biological assays.

Multiplex polymerase chain reaction (PCR) is an extension of the standard PCR protocol in which primers for multiple DNA loci are pooled together within a single reaction tube, enabling simultaneous sequence amplification, thus reducing costs and saving time. Potential cost saving and throughput improvements directly depend on the level of multiplexing achieved. Designing reliable and highly multiplexed assays is challenging because primers that are pooled together in a single reaction tube may cross-hybridize, though this can be addressed either by modifying the choice of primers for one or more amplicons, or by altering the way in which DNA loci are partitioned into separate reaction tubes. In this paper, we introduce a new graph formalism called a multi-node graph, and describe its application to the analysis of multiplex PCR scalability. We show, using random multi-node graphs that the scalability of multiplex PCR is constrained by a phase transition, suggesting fundamental limits on efforts to improve the cost-effectiveness and throughput of standard multiplex PCR assays. In particular, we show that when the multiplexing level of the reaction tubes is roughly theta(log (sn)) (where s is the number of primer pair candidates per locus and n is the number of loci to be amplified), then with very high probability we can 'cover' all loci with a valid assignment to one of the tubes in the assay. However, when the multiplexing level of the tube exceeds these bounds, there is no possible cover and moreover the size of the cover drops dramatically. Simulations using a simple greedy algorithm on real DNA data also confirm the presence of this phase transition. Our theoretical results suggest, however, that the resulting phase transition is a fundamental characteristic of the problem, implying intrinsic limits on the development of future assay design algorithms.     

Effects of NaCl on surface properties,chlorophyll fluorescence and light remission,and cellular compounds of <Emphasis Type="Italic">Grewia tenax</Emphasis> (Forssk.) Fiori and <Emphasis Type="Italic">Tamarindus indica</Emphasis> L. leaves

Seedlings of the salt-tolerant plant grewia [Grewia tenax (Forssk.) Fiori] and the moderately salt-tolerant tamarind (Tamarindus indica L.) were grown under controlled conditions and treated daily with NaCl solutions to investigate mechanisms of tolerance to salinity. Leaf micromorphology, cuticular wax load, chlorophyll fluorescence and light remission, as well as antioxidative potential were evaluated. As confirmed by energy-dispersive X-ray microanalysis in both species, absorption of sodium and chlorine increased with rising NaCl concentration in the treatment solution. In parallel, accumulation of calcium in grewia leaves was strongly reduced, leading to less crystals of calcium oxalate in leaf tissue. In grewia the cuticular wax load, chlorophyll content, and electron transport rate (ETR) were significantly reduced by comparatively low NaCl concentrations. In tamarind, in contrast, wax load and ETR were not significantly affected, while the decrease of chlorophyll content was less pronounced. Measurements of the antioxidative capacity and the imbalance between values of lipophilic and hydrophilic extracts at different NaCl concentrations confirmed that grewia is more salt tolerant than tamarind. This higher tolerance degree seemed to be associated with grewias’ more efficient scavenging of free radicals and the regulation of the antioxidative potential in lipophilic and hydrophilic extracts.     

Molecular keys unlock the mysteries of variable survival responses of blue crabs to hypoxia

Hypoxia is a major stressor in coastal ecosystems, yet generalizing its impacts on fish and shellfish populations across hypoxic events is difficult due to variability among individuals in their history of exposure to hypoxia and related abiotic variables, and subsequent behavioral and survival responses. Although aquatic animals have diverse physiological responses to cope with hypoxia, we know little about how inter-individual variation in physiological state affects survival and behavioral decisions under hypoxic conditions. Laboratory experiments coupled with molecular techniques determined how extrinsic factors (e.g., water body and temperature) and respiratory physiology (hemocyanin concentration and structure) affected survival and behavior of adult blue crabs (Callinectes sapidus) exposed to different levels of hypoxia over a 30-h time period. Nearly 100% of crabs survived the 1.3 mg dissolved oxygen (DO) l^?1 treatment (18.4% air saturation), suggesting that adult blue crabs are tolerant of severe hypoxia. Probability of survival decreased with increasing hypoxic exposure time, lower DO, and increasing temperature. Individual-level differences in survival correlated with water body and crab size. Crabs collected from the oligo/mesohaline and hypoxic Neuse River Estuary (NRE), North Carolina, USA survived hypoxic exposures longer than crabs from the euhaline and normoxic Bogue and Back Sounds, North Carolina. Furthermore, small NRE crabs survived longer than large NRE crabs. Hemocyanin (Hcy) concentration did not explain these individual-level differences, however, hypoxia-tolerant crabs had Hcy structures indicative of a high-O₂-affinity form of Hcy, suggesting Hcy “quality” (i.e., structure) may be more important for hypoxia survival than Hcy “quantity” (i.e., concentration). The geographic differences in survival we observed also highlight the importance of carefully selecting experimental animals when planning to extrapolate results to the population level.     

Approximate Maximum Parsimony and Ancestral Maximum Likelihood

We explore the maximum parsimony (MP) and ancestral maximum likelihood (AML) criteria in phylogenetic tree reconstruction. Both problems are NP-hard, so we seek approximate solutions. We formulate the two problems as Steiner tree problems under appropriate distances. The gist of our approach is the succinct characterization of Steiner trees for a small number of leaves for the two distances. This enables the use of known Steiner tree approximation algorithms. The approach leads to a 16/9 approximation ratio for AML and asymptotically to a 1.55 approximation ratio for MP.     

The relationship between the golden spiny mouse circadian system and its diurnal activity: an experimental field enclosures and laboratory study

Examples of animals that switch activity times between nocturnality and diurnality in nature are relatively infrequent. Furthermore, the mechanism for switching activity time is not clear: does a complete inversion of the circadian system occur in conjunction with activity pattern? Are there switching centers downstream from the internal clock that interpret the clock differently? Or does the switch reflect a masking effect? Answering these key questions may shed light on the mechanisms regulating activity patterns and their evolution. The golden spiny mouse (Acomys russatus) can switch between nocturnal and diurnal activity. This study investigated the relationship between its internal circadian clock and its diurnal activity pattern observed in the field. The goal is to understand the mechanisms underlying species rhythm shifts in order to gain insight into the evolution of activity patterns. All golden spiny mice had opposite activity patterns in the field than those under controlled continuous dark conditions in the laboratory. Activity and body temperature patterns in the field were diurnal, while in the laboratory all individuals immediately showed a free-running rhythm starting with a nocturnal pattern. No phase transients were found toward the preferred nocturnal activity pattern, as would be expected in the case of true entrainment. Moreover, the fact that the free-running activity patterns began from the individuals' subjective night suggests that golden spiny mice are nocturnal and that their diurnality in their natural habitat in the field results from a change that is downstream to the internal clock or reflects a masking effect.