PubMed Health⌕ Search

Biomedical subjects

S R Pai

Publications and source records attributed to S R Pai.

At least 19 recordsLinked to original sources
PubMed2000

Identification of viable and non-viable Mycobacterium tuberculosis in mouse organs by directed RT-PCR for antigen 85B mRNA.

Mycobacterium tuberculosis (MTB) the causative organism of tuberculosis can remain dormant as a non-culturable organism, reactivate and cause disease in man and animals. There is a need for proof of viability of such organisms in order to understand the process of reactivation. PCR for bacterial DNA cannot distinguish between viable and non-viable bacilli. We have tested a previously described two tube directed reverse transcriptase polymerase chain reaction (RT-PCR) for the detection of mRNA of antigen 85B (Ag85B) of MTB that can distinguish between viable and non-viable organisms. Using a set of external and internal primers for Ag85B, a cDNA amplified product (216 bp) was seen among simulated samples containing only viable cfus at a sensitivity of >10 and <100 cfu/ml. Eucaryotic DNA rich normal mouse lung homogenate did not interfere among these samples. The method amplified the 216 bp product also among cfu positive tissues of naturally infected mice. Finally, in a mouse model of dormancy, direct RT-PCR detected a signal among multiple tissues that were negative for cfus and hence non-culturable. Ag85B is abundantly secreted by MTB and hyper-expressed under stress conditions. Thus the method to identify its mRNA message may be useful to detect viable but dormant bacteria.

Acyltransferases↗
PubMed1992

Catabolic pools in Escherichia coli.

Methods are described for measuring soluble pool magnitudes in steady-state or exponentially growing cultures, and for distinguishing between anabolic, catabolic and total metabolic pools within cells. These methods were applied to the measurement of pool magnitudes for several amino acids and other precursors in Escherichia coli THU. Our results support the independence of the magnitudes of total metabolic pools and growth rate in steady-state cultures. Our results also show that the total metabolic pool size is much larger than previously published estimates, which failed to include the contribution of catabolic pools. The average value of the total soluble material in exponential-phase cells is estimated to be 8 to 9% of the cell dry mass; pool values could be almost twice as large during midcycle because of the known increase in the magnitudes of protein and RNA precursor pool during the cell cycle.

Amino Acids↗
PubMed1992

Characterization of monoclonal antibodies to the outer membrane protein (OmpD) of Salmonella typhimurium.

A panel of monoclonal antibodies, seven against the trimeric and seven against the monomeric forms to outer membrane protein D (OmpD) of Salmonella typhimurium were produced. The specificities of these monoclonal antibodies for the porin proteins of S. typhimurium and their cross-reactions with Salmonella porins OmpC and OmpF were determined by Western immunoblotting and enzyme-linked immunosorbent assay. We observed that OmpD shared more epitopes and had greater structural similarity with OmpC than with OmpF.

Animals↗
PubMed1992

Growth of HeLa S3 cells cotransfected with plasmids containing a c-fos gene under the control of the SV40 promoter complex, pRSVcat, and G418 resistance.

HeLa S3 cells, which have been fractionated into sequential and synchronous cell cycle phase-specific fractions, express c-fos at twice the basal levels in the earliest part of G1 phase. To determine whether this peak in c-fos synthesis has regulatory significance, a DNA construct was prepared which contained the human c-fos gene under the transcriptional control of the SV40 promoter complex. The pc-fos(human)-1 gene (9 kilobases) was inserted into the eukaryotic expression vector pSG5 (4.076 kilobases) at the EcoRI site. Electroporation with an exponentially decaying pulse was employed to cotransfect this construct into HeLa S3 cells along with the plasmids pRSVcat and the neomycin-resistance plasmid pF beta fos3' neo. The level of transient expression of each plasmid was determined. Transfection efficiency was determined as percentage fluorescent cells by measurement of immunofluorescence with a chloramphenicol acetyltransferase (CAT) antibody. Efficiency of transfection ranged up to approximately 5% of the cells. Transfected cells were selected on the basis of resistance to Geneticin (G418) at 400 micrograms/mL. CAT fluorescence and Geneticin resistance were employed to select permanently transformed cell lines. Compared with exponentially growing cells, successfully transfected cell lines expressed more than twice the level of c-fos mRNA as determined by dot-blot analysis and 16 times more of the 62-kilodalton c-fos protein as determined by Western blot analysis. As all cells in the population were not stable c-fos transfectants, this value is likely to be an underestimate of the overexpression level. In addition, expression was under the control of a strong serum induction insensitive promoter, unlike the native c-fos promoter.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetyltransferases↗
PubMed1991

The phosphate pool in Escherichia coli THU.

The phosphate pool of Escherichia coli was determined as a fraction of the total cell phosphate. This relative pool size was found to be essentially independent of cell age.

Escherichia coli↗
PubMed1991

Quick and high-efficiency electroelution of nucleic acid fragments.

We describe a quick, simple, and inexpensive technique for the electroelution of nucleic acid fragments that provides a high yield of DNA by using common laboratory components. The quantity of buffer used for the recovery of nucleic acid fragments is relatively small, the quality of the DNA recovered is relatively high, and more than one electroelution can be carried out at the same time.

Chemical Precipitation↗
PubMed1990

Response of lymphoid cells to thymic hormonal factors isolated from mouse thymus.

Thymic hormonal factors were isolated from mouse thymus by two methods. (1) Thymic cytosols in phosphate buffer saline were filtered through Sephadex G100 with 0.1 M NH4HCO3 (pH 8.0) as buffer and the protein peaks were collected. (2) Protein having thymosin activity (F5) was isolated from thymic cytosols after heat inactivation, salt fractionation and desalting on Sephadex G25. Molecular weights of all the proteins were determined on SDS-PAGE. Biological activity of thymic proteins was studied by in vitro and in vivo assays, using synthetic thymosin alpha 1 as the standard. Thymocytes treated with different thymic proteins showed maximum stimulation at 16 h of incubation period. Preincubation of the thymocytes with the thymic proteins and subsequent incubation with Con-A decreased the stimulation index. Incubation of spleen lymphocytes with thymic proteins increased the percentage of Tdt+ cells. The antitumor effects of thymic proteins carried out on animals having leukemia, showed statistically significant results. Clinically however, the antitumor effects of the thymic proteins alone and in combination chemotherapy were negligible at 1 mg/kg body weight dose level.

Animals↗
PubMed1989

Activation of amino acid transport during steady-state growth of Escherichia coli.

Accumulation of amino acids in exponentially increasing cultures of Escherichia coli was linear, supporting the interpretation that the biphasic response observed when cultures grew without these acids reflects a transient perturbation in accumulation. Rates of accumulation of glutamine, histidine and glycine were compared in steady-state and non-steady-state cultures. Their uptake rates were markedly enhanced in steady-state cultures at low exogenous concentrations, 10 microM or less. The results support the activation of amino acid transport systems by low concentrations of the particular amino acid present during growth. This activation was decreased when exogenous concentrations of the amino acid were markedly increased or when cells were washed free of the amino acid. Upon readdition of the amino acid after washing, recovery of enhanced transport required several generations, supporting a process of recovery other than enzymatic induction. The observation of amino acid enhancement of transport for eight other amino acids examined in steady-state culture suggests that this enhancement is a common process.

Amino Acids↗
PubMed1988

Variation in precursor pool size during the division cycle of Escherichia coli: further evidence for linear cell growth.

The magnitudes of several pools of radioactively labeled precursors for RNA and protein synthesis were determined as a function of cell age during the division cycle of Escherichia coli 15 THU. Uracil, histidine, and methionine pools increased from low initial values for cells at birth to maxima during midcycle and then subsided again. These pools were small or nonexistent at the beginning and the end of the cycle, and their average values during the cycle were less than 4% of the total cellular radioactivity. The results are consistent with a linear pattern of growth for cells during the division cycle and provide strong evidence against exponential or bilinear growth of E. coli cells.

Bacterial Proteins↗
PubMed1985

Acceleration of neoplasms of reticular tissue with syngeneic thymic graft under renal capsule and gamma radiation.

Weanling ICRC female mice were given syngeneic neonatal thymus graft under renal capsule and two weeks later whole-body gamma-radiation (group ITR). The findings on ITR were compared with three controls--untreated (C), thymus grafted (IT) and irradiated (IR). In ITR group 15/15 developed generalized neoplasms of reticular tissue at 5 to 8 months and single mammary tumor at 7 months. In IR group 12/13, 21/25 in IT and 5/23 in C had neoplastic changes at 7-month onwards. Microscopically these neoplasms had diffuse or focal proliferation of modified reticulum cells, histiocytes, lymphocytes and giant cells. Localized thymic tumors showed either the same picture or prominence of small lymphocytes. Mammary tumors developed in 12/25 IT, and 8/25 in C and nil in IR.

Animals↗
PubMed1984

Enhancement of leukemia and mammary tumor development in ICRC mice with subcapsular graft of neonatal thymus.

Weanling ICRC mice were given a syngeneic graft of neonatal thymus. The control (C) and experimental mice (IT) were observed for their natural death. Neoplasms of the reticular tissue developed in 80% (19/23) mice of IT group and 21% (5/23) in control. Mammary tumors developed in 52% (12/23) in IT group and 35% (8/23) in controls. Cell of origin of the leukemia was reticular in 9/23, lymphocytic in 7/23 and mixed lesion in 3/23 in IT group; while in controls it was 2/23 and 3/23, respectively and mixed lesion was absent. All mammary tumors were of adenocarcinoma series.

Adenocarcinoma↗
PubMed1981

Long-term feeding study in C17 mice administered saccharin coated betel nut and 1,4-dinitrosopiperazine in combination.

The observations on the effect of 3 agents--1,4-dinitrosopiperazine, betel nut and saccharin fed to C17 mice in combination is presented in this report. A total of 119 inbred mice of both sexes were put on long-term feeding trials. Group I consisted of 34 mice given a standard diet; group II of 32 mice fed an experimental diet containing saccharin coated betel nut powder at 10% concentration; group III of 29 mice given 0.2 ml aqueous solution of 0.1% 1,4-dinitrosopiperazine by intubation daily and group IV of 24 mice fed a combination of the experimental diet together with intubation of 1,4-dinitrosopiperazine. Feeding was continued for 40 weeks at which time all mice were given a standard diet and water ad libitum, and then observed for their full life-span. The commonest neoplasm found was squamous cell carcinoma of the forestomach in groups III and IV. Male mice were more susceptible to the treatment than female mice. In female mice reticular cell neoplasm--Type A of the uterus was the commonest tumour and was more common in group III than in group IV. The diet of saccharin coated betel nut failed to potentiate the carcinogenicity of 1,4-dinitrosopiperazine.

Animals↗