Manipulation of chromosomal DNA and localization of enzymatic activity

Observation of reaction among a few molecules will contribute to obtain profitably detailed information in biological fields such as molecular biology, which cannot be obtained from an in vitro system containing a large amount of molecules. This paper describes manipulation of a yeast chromosomal DNA and localization of enzymatic activity on a single or a few DNA molecules, as follows. (1) Globule structure in DNA molecules suppressed fragmentation of long DNA molecules due to sharing stress. It was demonstrated experimentally that the DNAs in globule state were tough for sharing stress. It means that globule formation enables mechanical manipulation of a long DNA molecule. We could make a successful recovery of a single globular chromosomal DNA into a glass capillary in a solution. (2) Using local area temperature control technique, activity of restriction enzyme could be activated only in a localized area by raising a local temperature. Then chromosomal DNAs were digested about 10 /spl mu/m around the laser focal spot on their DNAs. These single molecule technologies may contribute to several fields, especially biotechnology and genome analysis.