BACKGROUND & OBJECTIVES mRNA is more rapidly destroyed in cells than rRNA or genomic DNA, an assay targeting bacterial mRNA would provide a better guide to mycobacterial viability than amplification tests directed at DNA or rRNA targets. This study was carried out to standardize reverse transcriptase PCR (RT-PCR) targeting 85B gene for the rapid detection of viable Mycobacterium tuberculosis fr...

Huang, Ren-Qi, Zhenglan Chen, and Glenn H. Dillon. Molecular basis for modulation of recombinant 1 2 2 GABAA receptors by protons. J Neurophysiol 92: 883–894, 2004. First published March 17, 2004; 10.1152/jn.01040.2003. We have previously shown that extracellular protons inhibit recombinant and native GABAA receptors. In this report, we studied the site(s) and mechanism by which protons modulat...

The alkynyl phosphate ester, 1-hexynyl diethyl phosphate (I), is a mechanism-based inhibitor of phosphotriesterase. It has been previously determined that a histidine residue in the wild-type phosphotriesterase is covalently modified by this compound. In order to identify which of the seven histidine residues in the native enzyme are required for inactivation, the kinetic properties of phosphot...

There have been described a wide variety of methods for the preparation of total RNA from eucaryotic cells (1, 2). These methods are mainly designed for the isolation of RNA from relatively large amounts of cells. In a situation of screening a large number of cell clones, minimized cell cultivation effort and preparation time for total RNA isolation will be of great advantage. We developed a me...

Trichomoniasis, caused by the colonization of Trichomonas vaginalis in the urogenital tract, is one of the most prevalent sexually transmitted human diseases. Efforts to extract high-quality nucleic acids for studies have often been hampered by the potent nuclease activities released upon lysis of T. vaginalis (1–3,6,7). Protocols to isolate DNA from T. vaginalis, such as the guanidinium-HCl me...

The nuclear run-off transcription assay is currently the most sensitive technique to measure the in situ transcription of specific genes. It provides information on the synthesis of a specific gene that occurs as a function of cell state and is unaffected by potentially confounding posttranscriptional events such as RNA processing, transport, editing, and mRNA degradation (2). Briefly, this ass...

ii ACKNOWLEDGMENTS I have gained a vast amount of skill and knowledge throughout my Master of Science study at Louisiana State University. Therefore, I would like to first thank Dr. James V. Moroney, my major professor, for his patience and guidance and also Dr. C. Larkin, my committee members, for their help, guidance and time. I have studied in an environment surrounded by people who have rat...

Estimations of RNA abundance in solid tissues by nuclease protection assays (1, 2) are difficult, because of problems attaining efficient RNA extraction and because of RNA instability during molecular hybridization or after denaturation of nuclease-treated, probe:target hybrids. A method to overcome these difficulties in the case of tissue cultured cells was published (3) which conducted molecu...