A novel method for the two-dimensional analysis of proteins. |
| |
Authors: | K S Bloom J N Anderson |
| |
Institution: | Biology Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37830 USA |
| |
Abstract: | Nuclei from hamster embryo fibroblasts treated with radioactive benzo(a)pyrene were lysed in 6 m guanidine, and nuclear macromolecules were separated by isopycnic centrifugation in Cs2SO4. Control experiments showed that cross-contamination of the RNA, DNA, and protein fractions was less than 2% of the total recovery of each macromolecular class. When compared to previous techniques utilizing phenol extraction, similar specific activities of bound hydrocarbon (pmol benzoa]pyrene/mg protein or nucleic acid) were obtained. However, overall recoveries of macromolecular components were higher with the present method. In addition, recovery of undegraded histones in the density gradient preparation of nuclear protein was demonstrated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and recovery of native DNA was demonstrated by thermal denaturation studies. Although developed specifically for work with carcinogenic hydrocarbons, the Cs2SO4 technique should be generally useful in cases where it is necessary to prepare all three classes of macromolecules from one batch of nuclei. |
| |
Keywords: | |
本文献已被 ScienceDirect 等数据库收录! |
|