Computational SNP Analysis: Current Approaches and Future Prospects

The computational approaches in determining disease-associated Non-synonymous single nucleotide polymorphisms (nsSNPs) have evolved very rapidly. Large number of deleterious and disease-associated nsSNP detection tools have been developed in last decade showing high prediction reliability. Despite of all these highly efficient tools, we still lack the accuracy level in determining the genotype–phenotype association of predicted nsSNPs. Furthermore, there are enormous questions that are yet to be computationally compiled before we might talk about the prediction accuracy. Earlier we have incorporated molecular dynamics simulation approaches to foster the accuracy level of computational nsSNP analysis roadmap, which further helped us to determine the changes in the protein phenotype associated with the computationally predicted disease-associated mutation. Here we have discussed on the present scenario of computational nsSNP characterization technique and some of the questions that are crucial for the proper understanding of pathogenicity level for any disease associated mutations.     

Evaluating the Performance of Time-Gated Live-Cell Microscopy with Lanthanide Probes

Probes and biosensors that incorporate luminescent Tb(III) or Eu(III) complexes are promising for cellular imaging because time-gated microscopes can detect their long-lifetime (approximately milliseconds) emission without interference from short-lifetime (approximately nanoseconds) fluorescence background. Moreover, the discrete, narrow emission bands of Tb(III) complexes make them uniquely suited for multiplexed imaging applications because they can serve as Förster resonance energy transfer (FRET) donors to two or more differently colored acceptors. However, lanthanide complexes have low photon emission rates that can limit the image signal/noise ratio, which has a square-root dependence on photon counts. This work describes the performance of a wide-field, time-gated microscope with respect to its ability to image Tb(III) luminescence and Tb(III)-mediated FRET in cultured mammalian cells. The system employed a UV-emitting LED for low-power, pulsed excitation and an intensified CCD camera for gated detection. Exposure times of ∼1 s were needed to collect 5–25 photons per pixel from cells that contained micromolar concentrations of a Tb(III) complex. The observed photon counts matched those predicted by a theoretical model that incorporated the photophysical properties of the Tb(III) probe and the instrument’s light-collection characteristics. Despite low photon counts, images of Tb(III)/green fluorescent protein FRET with a signal/noise ratio ≥ 7 were acquired, and a 90% change in the ratiometric FRET signal was measured. This study shows that the sensitivity and precision of lanthanide-based cellular microscopy can approach that of conventional FRET microscopy with fluorescent proteins. The results should encourage further development of lanthanide biosensors that can measure analyte concentration, enzyme activation, and protein-protein interactions in live cells.     

In vivo detection of venous sinus distension due to intracranial hypotension in small animal using pulsed‐laser‐diode photoacoustic tomography

Intracranial hypotension (IH) is a pathophysiological condition of reduced intracranial pressure caused by low cerebrospinal fluid (CSF) volume due to dural injuries from lumbar puncture, surgery, or trauma. Understanding the prognosis of IH in small animal models is important to gain insights on the complications associated with it such as orthostatic headache, cerebral venous thrombosis, coma, and so forth. Photoacoustic tomography (PAT) offers a novel and cost‐effective way to perceive and detect IH in small animal models. In this study, a pulsed laser diode (PLD)‐based PAT imaging system was used to examine the changes in the venous sinuses of the rat brain due to IH, induced through CSF extraction. After the CSF extraction, an increase in the sagittal sinus area by ~30% and width by 40% ± 5% was observed. These results provide supportive evidence that the PLD‐PAT can be employed for detecting changes in sagittal sinus due to IH in rat model.     

Sodium nitroprusside enhances callus induction and shoot regeneration in high value medicinal plant Canscora diffusa

Plant Cell, Tissue and Organ Culture (PCTOC) - In different countries, mostly in EU, rules for strawberry nursery propagation impose the use of micropropagation only to produce stock virus free...     

Establishing catalytic activity on an artificial (βα)8-barrel protein designed from identical half-barrels

It has been postulated that the ubiquitous (βα)₈-barrel enzyme fold has evolved by duplication and fusion of an ancestral (βα)₄-half-barrel. We have previously reconstructed this process in the laboratory by fusing two copies of the C-terminal half-barrel HisF-C of imidazole glycerol phosphate synthase (HisF). The resulting construct HisF-CC was stepwise stabilized to Sym1 and Sym2, which are extremely robust but catalytically inert proteins. Here, we report on the generation of a circular permutant of Sym2 and the establishment of a sugar isomerization reaction on its scaffold. Our results demonstrate that duplication and mutagenesis of (βα)₄-half-barrels can readily lead to a stable and catalytically active (βα)₈-barrel enzyme.