High frequency in vitro regeneration system for conservation of Coleus forskohlii: a threatened medicinal herb

Present study reports a high frequency regeneration system for in vitro propagation and conservation of an important and threatened medicinal herb Coleus forskohlii (Briq.). Shoot multiplication has been achieved through axillary bud development and direct adventitious shoot formation in nodal explants on Murashige and Skoog (MS) medium containing 6-benzyladenine (BA) (5 μM). Further shoot multiplication was recorded up to third subculture on MS medium containing BA (5 μM) in combination with 1-naphthleneacetic acid (NAA) (0.1 μM). Excised microshoots on transfer to root induction medium consisting of half-strength MS medium (1/2 MS) alone as well as in combination with various auxins, resulted in varied rooting pattern in terms of number, length, and type of roots. Rooted microshoots were acclimatized successfully in earthen pots containing garden manure, garden soil, and sand (1:2:1) as potting mix with survival rate of 70 %. Acclimatized plantlets were studied for the amount of chlorophyll and carotenoid content as well as the net photosynthetic rate (P_N) during subsequent weeks of transfer to ex vitro condition. Histological studies revealed the direct origin and development of shoot buds from basal swollen cut end of nodal explants.     

Distribution of beta-globin haplotypes among the tribes of southern Gujarat,India

The present study was carried out in Indo-European speaking tribal population groups of southern Gujarat (India) to elucidate the allelic and haplotypic content of β-globin system in individuals with HbAA genotypes. 6 neutral restriction sites of the β-globin system were analysed and various statistical parameters were estimated to draw meaningful interpretations. All the 6 sites were found to be polymorphic and most were in Hardy–Weinberg Equilibrium in the studied group. Haplotypes were constructed using two different combinations of the 6 restriction sites analysed. Analysis of the 5 sites revealed a set of three predominant haplotypes, ‘+−−−−’, ‘−++−+’ and ‘−+−++’; and haplotypes ‘+−−’, ‘++−’ and ‘+++’ were found to be the most frequent when the 3 sites were used to construct the haplotypes. Haplotypic heterozygosity levels (> 83%) observed in the present study group were comparable to those observed in African and Afro-American populations and greater than other world populations. All the ancestral haplotypes, +−−−−−, −++−+, −+−++ and −−−−+ were found in the study group. The distribution pattern of various haplotypes was consistent with the global pattern. The paucity of comparable data from other Indian populations restricted one from making interpretations about the study group's relationships with other Indian populations but the results were indicative of older population histories or experience of gene flow by the study group and their affinities with populations of southern India.     

Cell in situ zymography: an in vitro cytotechnology for localization of enzyme activity in cell culture

In situ zymography is a unique technique for detection and localization of enzyme?Csubstrate interactions majorly in histological sections. Substrate with quenched fluorogenic molecule is incorporated in gel over which tissue sections are mounted and then incubated in buffer. The enzymatic activity is observed in the form of fluorescent signal. With the advancements in the field of biological research, use of in vitro cell culture has become very popular and holds great significance in multiple fields including inflammation, cancer, stem cell biology and the still emerging 3-D cell cultures. The information on analysis of enzymatic activity in cell lines is inadequate presently. We propose a single-step methodology that is simple, sensitive, cost-effective, and functional to perform and study the ??in position?? activity of enzyme on substrate for in vitro cell cultures. Quantification of enzymatic activity to carry out comparative studies on cells has also been illustrated. This technique can be applied to a variety of enzyme classes including proteases, amylases, xylanases, and cellulases in cell cultures.     

In Silico Studies in Probing the Role of Kinetic and Structural Effects of Different Drugs for the Reactivation of Tabun-Inhibited AChE

We have examined the reactivation mechanism of the tabun-conjugated AChE with various drugs using density functional theory (DFT) and post-Hartree-Fock methods. The electronic environments and structural features of neutral oximes (deazapralidoxime and 3-hydroxy-2-pyridinealdoxime) and charged monopyridinium oxime (2-PAM) and bispyridinium oxime (Ortho-7) are different, hence their efficacy varies towards the reactivation process of tabun-conjugated AChE. The calculated potential energy surfaces suggest that a monopyridinium reactivator is less favorable for the reactivation of tabun-inhibited AChE compared to a bis-quaternary reactivator, which substantiates the experimental study. The rate determining barrier with neutral oximes was found to be ∼2.5 kcal/mol, which was ∼5.0 kcal/mol lower than charged oxime drugs such as Ortho-7. The structural analysis of the calculated geometries suggest that the charged oximes form strong O^…H and N^…H hydrogen bonding and C-H^…π non-bonding interaction with the tabun-inhibited enzyme to stabilize the reactant complex compared to separated reactants, which influences the activation barrier. The ability of neutral drugs to cross the blood-brain barrier was also found to be superior to charged antidotes, which corroborates the available experimental observations. The calculated activation barriers support the superiority of neutral oximes for the activation of tabun-inhibited AChE compared to charged oximes. However, they lack effective interactions with their peripheral sites. Docking studies revealed that the poor binding affinity of simple neutral oxime drugs such as 3-hydroxy-2-pyridinealdoxime inside the active-site gorge of AChE was significantly augmented with the addition of neutral peripheral units compared to conventional charged peripheral sites. The newly designed oxime drug 2 appears to be an attractive candidate as efficient antidote to kinetically and structurally reactivate the tabun-inhibited enzyme.     

Haptoglobin polymorphism among the tribal groups of southern Gujarat

BACKGROUND:

Gujarat is located at the western most point of the Indian subcontinent. Valsad and Surat districts are part of the ‘tribal belt’of Gujarat and constitute 29.1% of total tribal population of Gujarat. These tribal populations are a rich source of gaining insights in the patterns of genetic diversity and genetico-environmental disorders against the back drop of their ecological, historical and ethnographic aspects.

AIM:

The objectives were to find out a) the genetic diversity among the tribes of Gujarat with reference to haptoglobin (Hp) locus b) the relationship between Hp polymorphism and sickle cell anemia/trait.

MATERIALS AND METHODS:

431 individuals belonging to eight tribal groups were studied for Hp polymorphism using polyacrylamide disc gel electrophoresis (PAGE). Hb*S was screened by dithionate tube turbididy (DTT) test and confirmed using cellulose acetate membrane electrophoresis (CAME).

STATISTICAL ANALYSIS:

Allele frequency was calculated by direct gene counting method. Average heterozygosity and gene diversity were computed using software DISPAN. Analysis of molecular variance (AMOVA) was estimated using software ARLEQUIN version 3.1.

RESULTS AND CONCLUSIONS:

Pattern of allele frequency distribution showed preponderance of Hp² allele in all the eight tribal groups, which is in accordance with its frequency in different populations of Indian subcontinent. Total average heterozygosity (H_T) was found to be low (0.160) but the level of genetic differentiation (G_ST) was found to be moderately high (5.6%). AMOVA analysis indicated least among group variance between west and south Indian populations (-0.04%) indicating the affinities of the tribes of Gujarat with that of Dravidian speaking groups. Analysis of Hp phenotypes among sickle cell anemia/ trait individuals revealed a high frequency of Hp 0-0 phenotype (92.7%) among SS individuals as opposed to only 9.7% among AS individuals, reaffirming the selective advantage of HbAS state in relation to hemolytic disorders.     

Microbial synthetic biology for human therapeutics

The emerging field of synthetic biology holds tremendous potential for developing novel drugs to treat various human conditions. The current study discusses the scope of synthetic biology for human therapeutics via microbial approach. In this context, synthetic biology aims at designing, engineering and building new microbial synthetic cells that do not pre-exist in nature as well as re-engineer existing microbes for synthesis of therapeutic products. It is expected that the construction of novel microbial genetic circuitry for human therapeutics will greatly benefit from the data generated by ??omics?? approaches and multidisciplinary nature of synthetic biology. Development of novel antimicrobial drugs and vaccines by engineering microbial systems are a promising area of research in the field of synthetic biology for human theragnostics. Expression of plant based medicinal compounds in the microbial system using synthetic biology tools is another avenue dealt in the present study. Additionally, the study suggest that the traditional medicinal knowledge can do value addition for developing novel drugs in the microbial systems using synthetic biology tools. The presented work envisions the success of synthetic biology for human therapeutics via microbial approach in a holistic manner. Keeping this in view, various legal and socio-ethical concerns emerging from the use of synthetic biology via microbial approach such as patenting, biosafety and biosecurity issues have been touched upon in the later sections.     

Genomic Structure of the Immigrant Siddis of East Africa to Southern India: A Study of 20 Autosomal DNA Markers

The Siddis are a tribal group of African origin living in Karnataka, India. They have undergone considerable cultural change due to their proximity to neighboring population groups. To understand the biological consequences of these changes, we describe the genomic structure of the Siddis and the contribution from putative ancestral populations using 20 autosomal DNA markers. The distribution of Alu indel markers and a genetic distance analysis reveals their closer affinities with Africans. The levels of genomic diversity and heterozygosity are high in all the populations of southern India. Genetic admixture analysis reveals a predominant contribution from Africans, a lesser contribution from south Indians, and a slight one from Europeans. There is no evidence of gametic disequilibrium in the Siddis. The genetic homogeneity of the Siddis, in spite of its admixed origin, suggests the utility of this population for genetic epidemiological studies.