Science: Uncovering and Discovering

Carbonic anhydrase (CA) has been of interest recently for its potential application in CO2 sequestration. Several groups have attempted application of CA enzymes or whole cells for this purpose. (more…)

Carbonic anhydrase (CA; EC 4.2.1.1) is a zinc-containing metalloenzyme catalyzing the reversible hydration of CO2. It has been found in all kingdoms of life, including both prokaryotic and eukaryotic subdivisions. Phylogenetically, CA can be divided into three distinct classes (α, β, and γ) that have no sequence homology and are supposed to have evolved independently. CA is important in biological systems because the uncatalyzed interconversion between CO2 and HCO3- is slow around neutral pH. Its high efficiency catalysis is fundamental to many biological processes, such as metabolism, photosynthesis, respiration, pH homeostasis and ion transport. (more…)

As discussed in the previous blog, the major disadvantage of chemical cloud seeding is that it uses harmful chemicals which get deposited in aquatic and terrestrial habitats following rain fall. Here they can have undesirable toxic and ecological effects on biota including plants and human beings. Secondly, abiotic particles such as dust and silver iodide are good at collecting water at temperatures below -8°C, but biological particles seem to be the main active nuclei at higher temperatures. Hence, there is a growing interest in biological particles which can act as cloud condensation nuclei. (more…)

From nowhere, artificial cloud seeding suddenly found itself making the headlines during the recent Olympic event in China. Many came to know about this phenomenon for the first time. However, you will be surprised to know that it is a very old science. So we can definitely spare some time to know more about it. (more…)

A zygote inherits a haploid set of chromosomes from both parents. Hence, its phenotype is supposed to result from co-expression of both alleles received from the sperm and the ovum. However, in mammals, gametes do not contribute equal genetic functions to the embryo due to the phenomenon of genomic imprinting. (more…)

As discussed in last blog, genome sequencing has revolutionized several fields in biology, antibiotic target discovery being one of them. Antibiotic targets can be of 2 types:
1) Proteins those are essential for bacterial growth.
2) Proteins those are required for virulence.
Such proteins can be identified by mutating all the genes from a pathogen, one at a time, and observing for the desired phenotype. STM is still another approach (discussed in previous blog). All these approaches, however, are labour intensive. (more…)

Advent of genome sequencing has revolutionized biology to its base. Development in approaches towards vaccine design is a good example of it. Vaccines have come a long way since their discovery by Jenner, a time when even their mode of action was unknown. Major development in vaccination then came up with the advent of recombinant DNA technology, which allowed selection and production of pure immunogens from pathogenic organisms. (more…)

The basic yeast two-hybrid system has been a widely used approach for general protein-protein interaction studies (as discussed in previous blog). With time researchers have modified it in many ways for certain special proteins that are not amenable to the basic system. Some of the modifications can even detect protein-nucleic acid and protein-ligand interactions. (more…)

The basic yeast two-hybrid system is based on the fact that eukaryotic transcriptional activators consist of a DNA binding domain (BD) and an activation domain (AD). Both these domains can be separately fused to 2 different polypeptides X and Y. if X and Y interact with each other, they can bring together BD and AD. This interaction can be monitored by transcriptional activation of a suitable reporter gene. (more…)

Application of proteomic techniques like 2-D gel electrophoresis (2-D GE) directly to crude cell extracts leads to loss of several protein constituents. This is due to the necessity of generalizing conditions for accommodating majority of proteins, rather than using conditions for a specific class of proteins. The resolution also decreases due to the broad range of separation parameters used in such techniques. These shortcomings have led researchers to use prefractionation techniques for enrichment of a particular group of proteins before separation by 2-D GE. (more…)

With increasing availability of whole gnome sequences, a new challenge has arised, that of understanding the function of each gene product, such as role in disease progression. The traditional methods involving testing mutants individually in animal models cannot handle genome-wide information. This has led to development of high throughput signature tagged mutagenesis (STM) approach. (more…)

One of the major challenges in bacterial expression profiling is the requirement for large amount of total RNA. This has led to development of amplification methods that can decrease the amount of starting material required and at the same time maintain the relative proportion of different mRNAs constant. (more…)

In the previous blog extraction of bacterial mRNA for global gene expression analysis was discussed. The next step in the analysis involves synthesis of cDNA from mRNAs before hybridization to microarray reporters. In eukaryotes this is relatively easy as mRNA molecules are polyadenylated, allowing use of oligo(dT) primers. For prokaryotes random hexamers have to be used. However, this may lead to inclusion of undesirable mRNAs, say from tissue/cells of interacting partner in case of interaction studies. (more…)

Success of microarray technology depends largely on extraction of adequate quantities of intact RNA. The problem of RNA concentration compounds especially for the low abundance mRNA species. While studying virulence-associated bacterial genes, this is directly dependent on the number of bacteria recoverable from the infection model. (more…)

One of the most intriguing questions about X-inactivation is how two X chromosomes with similar sequences can have exactly opposite transcriptional state. This property was ultimately linked to a particular region, X-inactivation center (xic), on X chromosome. This linkage was deduced from the outcome of X-to-autosome translocations. Xs that retained xic could still undergo X-inactivation, while those that lose this region could not. (more…)

Heterochromatin consists of the darkly stained regions of chromosome in the interphase nucleus. It is commonly found near the nuclear membrane. Heterochromatic DNA is highly condensed and is frequently associated with centromeres, telomeres and satellite sequences. The genes in such regions are normally in silenced state. (more…)

In the last blog, mechanism of X chromosome silencing in mammals was discussed. It has several implications apart from the dosage compensation described earlier. During early stage of female development, both Xs are active. Since inactivation is random, some cells inactivate paternal X, while others inactivate maternal X. Once a chromosome is inactivated, the inactivation is maintained in all the descending cells. As inactivation is brought about early in development, the adult female is a random patchwork of clonally derived cells, each cone expressing the same X-linked alleles. (more…)

We know that sex of a mammal is determined by its sex chromosome content, XY being male and XX being female. Since Y chromosome carries only a few genes, it is reasonable to assume that females have double the number of X-linked genes as compared to males. Yet females synthesize the same amount of X-linked protein products as male cells. Clearly, some form of dosage compensation mechanism must exist to account for this. (more…)

Although oligonucleotide-bases arrays are gaining popularity, PCR product-based array is a much more developed and established technology. The algorithms for PCR primer design are more advanced than those for oligonucleotide design. (more…)

Microarrays have a limitation of requiring high concentration of target DNA for efficient hybridization. Microelectronic arrays overcome this through the use of electronics. They are based on application of direct current (positive) to individual microelectrodes beneath the test sites. This causes rapid concentration of negatively charged nucleic acids over selected locations. These molecules can then be hybridized to previously addressed and attached nucleic acids at the test sites.
Such electronic addressing allows improvement in both time and efficiency of hybridization process. Reversal of electric potential then allows rapid removal of unhybridized molecules. Thus, the system allows simultaneous analysis of a single sample at multiple test sites, with the provision of washing unbound sample at desired stringency.