Analysis of urine sediments after kidney transplantation

We applied multiparameter flow cytometry for the first time to the classification of cells and particles of urine sediments after kidney transplantation. After fluorescent staining with acridine orange, cells and particles were checked for their size and their peak intensity of green and red fluorescences, known to be related to the amounts of intracellular DNA and intracellular RNA, respectively. Urine sediments were tested daily during a period of up to three weeks. Occurrence and disappearance of certain classes of cells and particles were found to be similar among different patients during normal healing. The results from patients without rejection of the graft are suggested as baseline data, differences from which should be detectable with high sensitivity due to the large number of cells being examined.     

Yeast L double-stranded ribonucleic acid is synthesized during the G1 phase but not the S phase of the cell cycle.

The cytoplasm of Saccharomyces cerevisiae contains two major classes of protein-encapsulated double-stranded ribonucleic acids (dsRNA's), L and M. Replication of L and M dsRNA's was examined in cells arrested in the G1 phase by either alpha-factor, a yeast mating pheromone, or the restrictive temperature for a cell cycle mutant (cdc7). [3H]uracil was added during the arrest periods to cells prelabeled with [14C]uracil, and replication was monitored by determining the ratio of 3H/14C for purified dsRNA's. Like mitochondrial deoxyribonucleic acid, both L and M dsRNA's were synthesized in the G1 arrested cells. The replication of L dsRNA was also examined during the S phase, using cells synchronized in two different ways. Cells containing the cdc7 mutation, treated sequentially with alpha-factor and then the restrictive temperature, enter a synchronous S phase when transferred to permissive temperature. When cells entered the S phase, synthesis of L dsRNA ceased, and little or no synthesis was detected throughout the S phase. Synthesis of L dsRNA was also observed in G1 phase cells isolated from asynchronous cultures by velocity centrifugation. Again, synthesis ceased when cells entered the S phase. These results indicate that L dsRNA replication is under cell cycle control. The control differs from that of mitochondrial deoxyribonucleic acid, which replicates in all phases of the cell cycle, and from that of 2-micron DNA, a multiple-copy plasmid whose replication is confined to the S phase.     

Restriction analysis of the Streptomyces glaucescens genome by agarose gel electrophoresis

A method for the analysis of total DNA of Streptomyces glaucescens is described. The relevant steps are (a) extraction and purification of DNA, (b) restriction of DNA samples with type II restriction enzymes, (c) one dimensional separation of restriction fragments by agarose gel electrophoresis. A typical banding pattern was obtained for each wild type strain, independant of growth conditions or age of the culture. Mutant strains exhibited in most cases the same banding pattern as the parent wild type strain. Only in one specific mutant class a fragment of about 9 megadalton was missing.