| As the vital place where the essential genome functions and the process of hereditary takes place, the structure of the chromatin has been widely interested. Although various architectural models of the "30nm" chromatin fiber had been proposed based on different experimental materials and observations, they still remain controversial. We choose Hela cells as the experimental material, take the previous experimental method as a basis. Considering the characteristics of Hela cells, we do some improvement and innovation of the experimental method to make the program be more suitable for the study of this subject. Here, we extracted and purified "30nm" chromatin fibers from human HeLa S3cells and analyzed the effect of salt concentration on the appearance and compactness of the30nm chromatin fiber by electron microscopy. The program as:First, extracting and purifying chromatin fibers within the buffer closest to the intracellular environment; then, preparing electron microscopy samples and observing; at last, analyzing the experiment results basis on the biochemical test results and electron micrographs. Through the experiment, we found the following preliminary conclusions:1, the endogenous nuclease activities in the Hela S3cell could be kept low by carefully adjusting the ion concentrations (Na+, Ca2+, Mg2+) in the digestion buffer during the extraction of the "30nm" chromatin fibers;2, the salt (Na+) concentration was verified to have a big impact on the Hela S3chromatin fiber morphology-with the salt concentration increasing (from0mM Na+to100mM Na+), the chromatin fiber showed an increase in the extent of condensation;3, different length of chromatin fragments tend to different models which performances:short segment chromatin structer closer to the one-start model and long segment chromatin structer closer to the two-start model;4, Frozen electron microscopic observation of Hela cell chromatin fiber structure show some characteristics of two-start structure model. |
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