Nuclear Pore Complex Number and Distribution throughout the Saccharomyces cerevisiae Cell Cycle by Three-Dimensional Reconstruction from Electron Micrographs of Nuclear Envelopes

The number of nuclear pore complexes (NPCs) in individual nuclei of the yeast Saccharomyces cerevisiae was determined by computer-aided reconstruction of entire nuclei from electron micrographs of serially sectioned cells. Nuclei of 32 haploid cells at various points in the cell cycle were modeled and found to contain between 65 and 182 NPCs. Morphological markers, such as cell shape and nuclear shape, were used to determine the cell cycle stage of the cell being examined. NPC number was correlated with cell cycle stage to reveal that the number of NPCs increases steadily, beginning in G1-phase, suggesting that NPC assembly occurs continuously throughout the cell cycle. However, the accumulation of nuclear envelope observed during the cell cycle, indicated by nuclear surface area, is not continuous at the same rate, such that the density of NPCs per unit area of nuclear envelope peaks in apparent S-phase cells. Analysis of the nuclear envelope reconstructions also revealed no preferred NPC-to-NPC distance. However, NPCs were found in large clusters over regions of the nuclear envelope. Interestingly, clusters of NPCs were most pronounced in early mitotic nuclei and were found to be associated with the spindle pole bodies, but the functional significance of this association is unknown.

A two-year experience teaching computer literacy to first-year medical students using skill-based cohorts

Because it is widely accepted that providing information online will play a major role in both the teaching and practice of medicine in the near future, a short formal course of instruction in computer skills was proposed for the incoming class of students entering medical school at the State University of New York at Stony Brook. The syllabus was developed on the basis of a set of expected outcomes, which was accepted by the dean of medicine and the curriculum committee for classes beginning in the fall of 1997. Prior to their arrival, students were asked to complete a self-assessment survey designed to elucidate their initial skill base; the returned surveys showed students to have computer skills ranging from complete novice to that of a systems engineer. The classes were taught during the first three weeks of the semester to groups of students separated on the basis of their knowledge of and comfort with computers. Areas covered included computer basics, e-mail management, MEDLINE, and Internet search tools. Each student received seven hours of hands-on training followed by a test. The syllabus and emphasis of the classes were tailored to the initial skill base but the final test was given at the same level to all students. Student participation, test scores, and course evaluations indicated that this noncredit program was successful in achieving an acceptable level of comfort in using a computer for almost all of the student body.

Simultaneous assessment of loss of heterozygosity at multiple microsatellite loci using semi-automated fluorescence-based detection: subregional mapping of chromosome 4 in cervical carcinoma.

Detection of loss of heterozygosity (LOH) by comparison of normal and tumor genotypes using PCR-based microsatellite loci provides considerable advantages over traditional Southern blotting-based approaches. However, current methodologies are limited by several factors, including the numbers of loci that can be evaluated for LOH in a single experiment, the discrimination of true alleles versus "stutter bands," and the use of radionucleotides in detecting PCR products. Here we describe methods for high throughput simultaneous assessment of LOH at multiple loci in human tumors; these methods rely on the detection of amplified microsatellite loci by fluorescence-based DNA sequencing technology. Data generated by this approach are processed by several computer software programs that enable the automated linear quantitation and calculation of allelic ratios, allowing rapid ascertainment of LOH. As a test of this approach, genotypes at a series of loci on chromosome 4 were determined for 58 carcinomas of the uterine cervix. The results underscore the efficacy, sensitivity, and remarkable reproducibility of this approach to LOH detection and provide subchromosomal localization of two regions of chromosome 4 commonly altered in cervical tumors.