Genome sequencing of cells that live inside glass cages reveals their past history

One-quarter of the total primary production on earth is contributed by diatoms1. These are photosynthetic, unicellular algae with ornamented silica shells found in all aquatic and moist environments. They form the base of energy-efficient food webs that support all aquatic life forms. More than 250 genera of living diatoms, with as many as 100,000 species are known2. Fossil diatoms are known as early as the Cretaceous, 144–65 m.y. ago3. In India, deposits of diatoms occur in Rajasthan and are known as ‘multani mitti’. Multani mitti or Indian Fuller’s earth or diatomaceous earth as it is called in the West, is applied as a paste on the surface of the skin for 15–20 min and then washed-off. This leaves the skin feeling smooth, soft, moist and rejuvenated. Diatomaceous earth is now being used in the formulation of soaps, cleansing products, face powders and skincare preparations. Diatomaceous earth is a mineral material consisting mainly of siliceous fragments of various species of fossilized remains of diatoms.

Cloning and sequencing of winged bean (Psophocarpus tetragonolobus) basic agglutinin (WBA I): presence of second glycosylation site and its implications in quaternary structure

We report cloning of the DNA encoding winged bean basic agglutinin (WBA I). Using oligonucleotide primers corresponding to N- and C-termini of the mature lectin, the complete coding sequence for WBA I could be amplified from genomic DNA. DNA sequence determination by the chain termination method revealed the absence of any intervening sequences in the gene. The DNA deduced amino acid sequence of WBA I displayed some differences with its primary structure established previously by chemical means. Comparison of the sequence of WBA I with that of other legume lectins highlighted several interesting features, including the existence of the largest specificity determining loop which might account for its oligosaccharide-binding specificity and the presence of an additional N-glycosylation site. These data also throw some light on the relationship between the primary structure of the protein and its probable mode of dimerization.

Novel melody line identification algorithm for polyphonic MIDI music

The problem of automatic melody line identification in a MIDI file plays an important role towards taking QBH systems to the next level. We present here, a novel algorithm to identify the melody line in a polyphonic MIDI file. A note pruning and track/channel ranking method is used to identify the melody line. We use results from musicology to derive certain simple heuristics for the note pruning stage. This helps in the robustness of the algorithm, by way of discarding "spurious" notes. A ranking based on the melodic information in each track/channel enables us to choose the melody line accurately. Our algorithm makes no assumption about MIDI performer specific parameters, is simple and achieves an accuracy of 97% in identifying the melody line correctly. This algorithm is currently being used by us in a QBH system built in our lab.

Ultra high throughput sequencing excludes MDH1 as candidate gene for RP28-linked retinitis pigmentosa

Purpose: Mutations in IDH3B, an enzyme participating in the Krebs cycle, have recently been found to cause autosomal recessive retinitis pigmentosa (arRP). The MDH1 gene maps within the RP28 arRP linkage interval and encodes cytoplasmic malate dehydrogenase, an enzyme functionally related to IDH3B. As a proof of concept for candidate gene screening to be routinely performed by ultra high throughput sequencing (UHTs), we analyzed MDH1 in a patient from each of the two families described so far to show linkage between arRP and RP28. Methods: With genomic long-range PCR, we amplified all introns and exons of the MDH1 gene (23.4 kb). PCR products were then sequenced by short-read UHTs with no further processing. Computer-based mapping of the reads and mutation detection were performed by three independent software packages. Results: Despite the intrinsic complexity of human genome sequences, reads were easily mapped and analyzed, and all algorithms used provided the same results. The two patients were homozygous for all DNA variants identified in the region, which confirms previous linkage and homozygosity mapping results, but had different haplotypes, indicating genetic or allelic heterogeneity. None of the DNA changes detected could be associated with the disease. Conclusions: The MDH1 gene is not the cause of RP28-linked arRP. Our experimental strategy shows that long-range genomic PCR followed by UHTs provides an excellent system to perform a thorough screening of candidate genes for hereditary retinal degeneration.

Sequencing of T-superfamily conotoxins from Conus virgo: Pyroglutamic acid identification and disulfide arrangement by MALDI mass spectrometry

De novo mass spectrometric sequencing of two Conus peptides, Vi1359 and Vi1361, from the vermivorous cone snail Conus virgo, found off the southern Indian coast, is presented. The peptides, whose masses differ only by 2 Da, possess two disulfide bonds and an amidated C-terminus. Simple chemical modifications and enzymatic cleavage coupled with matrix assisted laser desorption ionization (MALDI) mass spectrometric analysis aided in establishing the sequences of Vi1359, ZCCITIPECCRI-NH2, and Vi1361, ZCCPTMPECCRI-NH2, Which differ only at residues 4 and 6 (Z = pyroglutamic acid). The presence of the pyroglutamyl residue at the N-terminus was unambiguously identified by chemical hydrolysis of the cyclic amide, followed by esterification. The presence of Ile residues in both the peptides was confirmed from high-energy collision induced dissociation (CID) studies, using the observation Of W-n- and d(n)-ions as a diagnostic. Differential cysteine labeling, in conjunction with MALDI-MS/MS, permitted establishment of disulfide connectivity in both peptides as Cys2-Cys9 and Cys3-Cys10. The cysteine pattern clearly reveals that the peptides belong to the class of T-superfamily conotoxins, in particular the T-1 superfamily.

Ultra high throughput sequencing excludes MDH1 as candidate gene for RP28-linked retinitis pigmentosa

Purpose: Mutations in IDH3B, an enzyme participating in the Krebs cycle, have recently been found to cause autosomal recessive retinitis pigmentosa (arRP). The MDH1 gene maps within the RP28 arRP linkage interval and encodes cytoplasmic malate dehydrogenase, an enzyme functionally related to IDH3B. As a proof of concept for candidate gene screening to be routinely performed by ultra high throughput sequencing (UHTs), we analyzed MDH1 in a patient from each of the two families described so far to show linkage between arRP and RP28. Methods: With genomic long-range PCR, we amplified all introns and exons of the MDH1 gene (23.4 kb). PCR products were then sequenced by short-read UHTs with no further processing. Computer-based mapping of the reads and mutation detection were performed by three independent software packages. Results: Despite the intrinsic complexity of human genome sequences, reads were easily mapped and analyzed, and all algorithms used provided the same results. The two patients were homozygous for all DNA variants identified in the region, which confirms previous linkage and homozygosity mapping results, but had different haplotypes, indicating genetic or allelic heterogeneity. None of the DNA changes detected could be associated with the disease.

A speech-music discriminator using HILN model based features

We propose a simple speech music discriminator that uses features based on HILN(Harmonics, Individual Lines and Noise) model. We have been able to test the strength of the feature set on a standard database of 66 files and get an accuracy of around 97%. We also have tested on sung queries and polyphonic music and have got very good results. The current algorithm is being used to discriminate between sung queries and played (using an instrument like flute) queries for a Query by Humming(QBH) system currently under development in the lab.

Finite data performance of MUSIC and minimum norm methods

In the direction of arrival (DOA) estimation problem, we encounter both finite data and insufficient knowledge of array characterization. It is therefore important to study how subspace-based methods perform in such conditions. We analyze the finite data performance of the multiple signal classification (MUSIC) and minimum norm (min. norm) methods in the presence of sensor gain and phase errors, and derive expressions for the mean square error (MSE) in the DOA estimates. These expressions are first derived assuming an arbitrary array and then simplified for the special case of an uniform linear array with isotropic sensors. When they are further simplified for the case of finite data only and sensor errors only, they reduce to the recent results given in [9-12]. Computer simulations are used to verify the closeness between the predicted and simulated values of the MSE.

Carnatic music analysis: Shadja, swara identification and rAga verification in AlApana using stochastic models

We analyze the AlApana of a Carnatic music piece without the prior knowledge of the singer or the rAga. AlApana is ameans to communicate to the audience, the flavor or the bhAva of the rAga through the permitted notes and its phrases. The input to our analysis is a recording of the vocal AlApana along with the accompanying instrument. The AdhAra shadja(base note) of the singer for that AlApana is estimated through a stochastic model of note frequencies. Based on the shadja, we identify the notes (swaras) used in the AlApana using a semi-continuous GMM. Using the probabilities of each note interval, we recognize swaras of the AlApana. For sampurNa rAgas, we can identify the possible rAga, based on the swaras. We have been able to achieve correct shadja identification, which is crucial to all further steps, in 88.8% of 55 AlApanas. Among them (48 AlApanas of 7 rAgas), we get 91.5% correct swara identification and 62.13% correct R (rAga) accuracy.

Protein Model Discrimination Using Mutational Sensitivity Derived from Deep Sequencing

A major bottleneck in protein structure prediction is the selection of correct models from a pool of decoys. Relative activities of similar to 1,200 individual single-site mutants in a saturation library of the bacterial toxin CcdB were estimated by determining their relative populations using deep sequencing. This phenotypic information was used to define an empirical score for each residue (Rank Score), which correlated with the residue depth, and identify active-site residues. Using these correlations, similar to 98% of correct models of CcdB (RMSD <= 4 angstrom) were identified from a large set of decoys. The model-discrimination methodology was further validated on eleven different monomeric proteins using simulated RankScore values. The methodology is also a rapid, accurate way to obtain relative activities of each mutant in a large pool and derive sequence-structure-function relationships without protein isolation or characterization. It can be applied to any system in which mutational effects can be monitored by a phenotypic readout.

Compressive sensing for music signals: comparison of transforms with coherent dictionaries

Compressive Sensing (CS) is a new sensing paradigm which permits sampling of a signal at its intrinsic information rate which could be much lower than Nyquist rate, while guaranteeing good quality reconstruction for signals sparse in a linear transform domain. We explore the application of CS formulation to music signals. Since music signals comprise of both tonal and transient nature, we examine several transforms such as discrete cosine transform (DCT), discrete wavelet transform (DWT), Fourier basis and also non-orthogonal warped transforms to explore the effectiveness of CS theory and the reconstruction algorithms. We show that for a given sparsity level, DCT, overcomplete, and warped Fourier dictionaries result in better reconstruction, and warped Fourier dictionary gives perceptually better reconstruction. “MUSHRA” test results show that a moderate quality reconstruction is possible with about half the Nyquist sampling.

Multiple parents crossover operators: A new approach removes the overlapping solutions for sequencing problems

Maintaining population diversity throughout generations of Genetic Algorithms (GAs) is key to avoid premature convergence. Redundant solutions is one cause for the decreasing population diversity. To prevent the negative effect of redundant solutions, we propose a framework that is based on the multi-parents crossover (MPX) operator embedded in GAs. Because MPX generates diversified chromosomes with good solution quality, when a pair of redundant solutions is found, we would generate a new offspring by using the MPX to replace the redundant chromosome. Three schemes of MPX will be examined and will be compared against some algorithms in literature when we solve the permutation flowshop scheduling problems, which is a strong NP-Hard sequencing problem. The results indicate that our approach significantly improves the solution quality. This study is useful for researchers who are trying to avoid premature convergence of evolutionary algorithms by solving the sequencing problems.

Genome sequencing unveils a novel sea enterotoxin-carrying PVL phage in staphylococcus aureus ST772 from India

Staphylococcus aureus is a major human pathogen, first recognized as a leading cause of hospital-acquired infections. Community-associated S. aureus (CA-SA) pose a greater threat due to increase in severity of infection and disease among children and healthy adults. CA-SA strains in India are genetically diverse, among which is the sequence type (ST) 772, which has now spread to Australia, Europe and Japan. Towards understanding the genetic characteristics of ST772, we obtained draft genome sequences of five relevant clinical isolates and studied the properties of their PVL-carrying prophages, whose presence is a defining hallmark of CA-SA. We show that this is a novel prophage, which carries the structural genes of the hlb-carrying prophage and includes the sea enterotoxin. This architecture probably emerged early within the ST772 lineage, at least in India. The sea gene, unique to ST772 PVL, despite having promoter sequence characteristics typical of low expression, appears to be highly expressed during early phase of growth in laboratory conditions. We speculate that this might be a consequence of its novel sequence context. The crippled nature of the hlb-converting prophage in ST772. suggests that widespread mobility of the sea enterotoxin might be a selective force behind its `transfer' to the PVL prophage. Wild type ST772 strains induced strong proliferative responses as well as high cytotoxic activity against neutrophils, likely mediated by superantigen SEA and the PVL toxin respectively. Both proliferation and cytotoxicity were markedly reduced in a cured ST772 strain indicating the impact of the phage on virulence. The presence of SEA alongside he genes for the immune system-modulating PVL toxin may contribute to the success and virulence of ST772.

Music and symbolic dynamics: the science behind an art

Music signals comprise of atomic notes drawn from a musical scale. The creation of musical sequences often involves splicing the notes in a constrained way resulting in aesthetically appealing patterns. We develop an approach for music signal representation based on symbolic dynamics by translating the lexicographic rules over a musical scale to constraints on a Markov chain. This source representation is useful for machine based music synthesis, in a way, similar to a musician producing original music. In order to mathematically quantify user listening experience, we study the correlation between the max-entropic rate of a musical scale and the subjective aesthetic component. We present our analysis with examples from the south Indian classical music system.