Effect of estrogen and its antagonist on the expression of arginine vasotocin (AVT) and its oxytocic-like receptor VT3 in the shell gland of Japanese quail, Coturnix coturnix japonica

Avian neurohypophysial hormone arginine vasotocin (AVT) is known to regulate shell gland contractility during oviposition. While studying the role of estrogen in the expression and regulation of AVT and its oxytocic-like receptor VT3, using in situ hybridization and immunohistochemistry, it was observed that the expression of AVT and its receptor was not detected in the shell gland of sexually immature Japanese quail. However, administration of estrogen to these birds not only stimulates the growth and activity (as assessed by increased mucosal fold length, total protein content and alkaline phosphatase level) of the shell gland but also upregulates the expression of AVT and VT3. Further, administration of estrogen antagonist tamoxifen to sexually mature bird shows opposite results. On the other hand, localization of ir-AVT, observed in the ovary of sexually mature bird, was not detected in the estrogen treated sexually immature quail. It is concluded that estrogen not only affects the growth and differentiation of avian oviduct, but also regulates the expression of shell gland AVT and its receptor VT3. Present findings suggest that the locally synthesized AVT acts in a paracrine way to upregulate VT3 receptor and thus facilitates the endocrine function of neurohypophysial AVT during oviposition.     

Production of schizophyllan using Schizophyllum commune NRCM

In the present work, four strains were screened for schizophyllan production, of which Schizophyllum commune NRCM was selected for further work. The fermentation was carried out for 168 h at 28+/-2 degrees C on an orbital shaker at 180 rpm. In the first step, one factor-at-a-time method was used to investigate the effect of media constituents such as carbon and nitrogen sources on schizophyllan production. Subsequently in the second step, concentration of the medium components was optimized using Response Surface Method (RSM). The yield increased from 3.25+/-0.72 g/l in the unoptimized media to 8.03+/-1.12 g/l in the medium optimized by RSM.     

Media optimization for the production of beta-carotene by Blakeslea trispora: a statistical approach

Blakeslea trispora (+) MTCC, Blakeslea trispora NRRL 2895 (+), Blakeslea trispora NRRL 2896 (-) as well as intraspecific mating of both the strain types have been studied for optimum production of beta-carotene. Intraspecific mating of both the strain types increased the yield of beta-carotene to a considerable level (98+/-2mg/l) as compared to wild strains. Effect of different media components such as carbon, nitrogen, and sulphates, and that of process variables such as pH and inoculum size on beta-carotene production by Blakeslea trispora in shake flask culture was investigated. One factor at-a-time method was employed for the optimization of media components. Response surface methodology (RSM) was further used to determine the optimum values of process variables for maximum beta-carotene production. The fit of the quadratic model was found to be significant. A significant increase in beta-carotene production (139+/-1mg/l) was achieved using RSM.     

Identification of genetically diverse genotypes for photoperiod insensitivity in soybean using RAPD markers

Most of the Indian soybean varieties were found to be highly sensitive to photoperiod, which limits their cultivation in only localized area. Identification of genetically diverse source of photoperiod insensitive would help to broaden the genetic base for this trait. Present study was undertaken with RAPD markers for genetic diversity estimation in 44 accessions of soybean differing in response to photoperiod sensitivity. The selected twenty-five RAPD primers produced a total of 199 amplicons, which generated 89.9 % polymorphism. The number of amplification products ranged from 2 to 13 for different primers. The polymorphism information content ranged from 0.0 for monomorphic loci to 0.5 with an average of 0.289. Genetic diversity between pairs of genotypes was 37.7% with a range of 3.9 to 71.6%. UPGMA cluster analysis placed all the accessions of soybean into four major clusters. No discernable geographical patterns were observed in clustering however; the smaller groups corresponded well with pedigree. Mantel’s test (r = 0.915) indicates very good fit for clustering pattern. Two genotypes, MACS 330 and 111/2/1939 made a very divergent group from other accessions of soybean and highly photoperiod insensitive that may be potential source for broadening the genetic base of soybean for this trait.Key words: Genetic diversity, Photoperiod, RAPD, Soybean, UPGMA     

Construction of a recF deletion mutant of Azotobacter vinelandii and its characterization

A deletion was engineered in the cloned recF gene by digestion with suitable restriction endonucleases and a tetracycline resistance gene cartridge was inserted. The mutation was subsequently transferred to the Azotobacter vinelandii chromosome by double cross-over under pressure of tetracycline selection. A recF recA mutant was also constructed in a similar manner. The mutations were found to be stable and mutation of the wild-type recF gene was confirmed by Southern blot hybridization. Both the mutants were UV sensitive and recombination deficient. Mutations in genes involved in nitrogen fixation in A. vinelandii are rather frequent and obtained comparatively easily despite of the presence of multiple identical chromosomes in A. vinelandii. It has been speculated that some kind of `homogenotization' process operates which is responsible for the `transmission' of mutation from one chromosome to all the chromosomes. This process is not affected by a mutation in recF or recA or in both recF and recA.     

Flotillin 2 is distinct from epidermal surface antigen (ESA) and is associated with filopodia formation.

ECS-1, a monoclonal antibody (MoAb) raised to cultured human keratinocytes, stains the intercellular glycocalyx with a pemphigus-like pattern and recognizes a 35-kDa epidermal surface antigen (ESA) on Western blotting of keratinocyte extracts. When ECS-1 MoAb was used to screen a keratinocyte expression library, a unique cDNA was identified that predicted a 42-kDa globular protein of unknown function. This putative ESA was conserved between mice and humans and was encoded by a gene on chromosome 17q11-12 in linkage with neurofibromin. Homology between the cDNA sequence has been reported with flotillin 1, a caveolae associated protein, as well as Reggie 1 and 2, neuronal proteins expressed during axonal regeneration present in activated GPI-anchored cell adhesion molecules in non-caveolar-associated micropatches. In order to determine whether the cDNA predicted protein and ECS-1 antigen were identical, we compared ECS-1 with the immunoreactivity of a new antibody raised to the cDNA fusion protein in epidermis and cultured cells. The cDNA fusion protein was expressed in bacteria and in cos cells with his, FLAG, and EGFP reporter tags and by stable transfection as an EGFP fusion protein. The fusion protein and native protein of 42 kDa were detected by the new antibody, but not by the original ECS-1. Thus, the ECS-1 antigen, ESA (35 kDa), is clearly distinct from the protein predicted by the cDNA (renamed flotillin 2). Stable transfection of ESA/flotillin 2 fusion protein in cos cells induced filopodia formation and changed epithelial cells to a neuronal appearance. Thus, the function of flotillin 2 may resemble that of the goldfish optic nerve neuronal regeneration proteins, Reggie 1 and 2.     

Effect of cadmium and zinc on antioxidant enzyme activity in the gastropod, Achatina fulica

Heavy metal stress results in the production of O(2)(.-), H(2)O(2) and (.)OH, which affect various cellular processes, mostly the functioning of membrane systems. Cells are normally protected against free oxyradicals by the operation of intricate antioxidant systems. The aim of the present work is to examine the effect of CdCl(2) and ZnSO(4) on antioxidative enzyme activity in the gastropod, Achatina fulica. The concentrations of antioxidant enzymes--superoxide dismutase (SOD), catalase (Cat) and glutathione peroxidase (GPx)--and nonenzymatic antioxidants--glutathione and vitamin-C--were found to be decreased in both digestive gland and kidney of the gastropod, Achatina fulica treated with individual concentrations of 0.5 ppm and 1ppm of CdCl(2) and ZnSO(4), compared to that of control animals. Based on the above study, it is evident that Achatina fulica can be used as a bioindicator to monitor the environmental heavy metal pollution.     

Cyclin-dependent kinase activating kinase/Cdk7 co-localizes with PKC-iota in human glioma cells

Cyclin-dependent kinase activating kinase (CAK) is a trimeric complex composed of cdk7, cyclin H and MAT1. CAK/cdk7 functions as a master cell cycle regulator by phosphorylating cyclin-dependent kinases for cell cycle progression. We have previously reported that protein kinase C-iota (PKC-iota) associates with CAK/cdk7. In this investigation, immunofluorescence confocal microscopy was used to provide further evidence for the co-localization of PKC-iota with CAK/cdk7. PKC-iota was labeled with Alexa Fluor 488 (green fluorescent dye) and CAK/cdk7 was labeled with Alexa Fluor 555 (red fluorescent dye). The fusion of the red and green fluorescent colors produced a yellow color, which was used to quantify co-localization of PKC-iota and CAK/cdk7. Confocal microscopy revealed the co-localization of PKC-iota with CAK/cdk7 in both the cytoplasm and nucleus of U-373 MG cells.