Segall 9733332 Many lines of evidence have shown that the structure of the E. coli and Salmonella bacterial chromosomes affects interactions with various enzyme complexes. Several probes of structure, including homologous recombination, transposition by Tn7 and Tn 10, and site-specific deletion mediated by the resolvase protein of the delta gamma transposon, have uncovered the presence of chromosomal "barriers". However, neither the nature of these barriers nor the basis of their action is known. Since these barriers are likely to affect all systems that interact with chromosomal DNA (repair, replication, and transcription functions as well as interactions between the chromosome and extrachromosomal elements like viruses and transposons), it is important to understand their nature and effects, and if and how they are regulated. The first stage is to unequivocally identify and define the barriers or structural elements. This research addresses this question in a systematic and more extensive manner than has been attempted in the past. The experiments are geared to understanding the three-dimensional structure of the chromosome within the bacterial cell, and how this structure is affected by some basic features of cell physiology. The probe that is being used is the site-specific recombination system of phage lambda, which has several features that make it a flexible and sensitive tool for the task at hand, and which will yield data complementary to that obtained by the probes used previously. The questions that are being addressed are: 1) Do all chromosomal sequences have equal access to each other? 2) Do circular extrachromosomal sequences (specifically plasmids) have equal access to all chromosomal locations? 3) Do loci that are easily accessible to extrachromosomal DNA but are poorly accessible to other chromosomal loci define regions that constrain the structure of the folded chromosome? One of the most limiting aspects of educating biology students today is hands-on laboratory traini ng not in a classical classroom context, but in a real-life research laboratory. Students with well-developed skills in this area are extremely sought-after, but too few of them have the opportunity to enter research labs and receive the necessary training in problem-solving skills and proficiency in lab methods, experimental design, and data analysis. To begin to correct this problem five summer research workshops are being formed for undergraduates where students will work in teams on different facets of a research problem. Students will be involved in the design, performance, and analysis of the experiments, and will learn modern molecular techniques. The workshops should benefit students most profoundly by exposing them to carrying out experiments under a more realistic setting than can be duplicated in larger laboratory courses in which students do set lab exercises. In addition, some of the experiments on chromosome structure will be adapted and introduced into one new and one ongoing laboratory course.
