Cadilla C L, Harp J, Flanagan J M, Olins A L, Olins D E
Nucleic Acids Res. 1986 Jan 24;14(2):823-41. doi: 10.1093/nar/14.2.823.
Euplotes eurystomus is a hypotrichous ciliate containing a transcriptionally active macronucleus (MAC) and a transcriptionally inactive micronucleus. Soluble MAC chromatin contains a normal complement of inner histones, an H1-like protein which is very sensitive to proteolysis, and a considerable proportion of non-histone proteins. A combination of N-Tosyl lysine chloromethyl ketone (TLCK) and PMSF was found to be most effective in preventing proteolysis. Microccocal nuclease digestion yielded an average nucleosome repeat length of 187 +/- 25 bp for soluble chromatin; and 190 +/- 9 bp for isolated macronuclei. Thermal denaturation profiles of MAC chromatin in 0.25 mM EDTA display two main transitions at about 76 and 83 degrees C, resembling the melting of soluble chicken erythrocyte chromatin. Circular dichroic spectra of MAC chromatin were compared to soluble chicken erythrocyte chromatin under the same ionic strength conditions and were found to be very similar. 2D chromatin/DNA agarose gel electrophoresis resulted in a diagonal line of DNA staining, which establishes a strict correlation between DNA and chromatin electrophoretic mobility.
宽口游仆虫是一种腹毛目纤毛虫，含有一个转录活跃的大核（MAC）和一个转录不活跃的小核。可溶性大核染色质包含正常的内部组蛋白成分、一种对蛋白水解非常敏感的H1样蛋白以及相当比例的非组蛋白。发现N-甲苯磺酰赖氨酸氯甲基酮（TLCK）和苯甲基磺酰氟（PMSF）的组合在防止蛋白水解方面最有效。微球菌核酸酶消化产生的可溶性染色质平均核小体重复长度为187±25 bp；分离的大核为190±9 bp。在0.25 mM乙二胺四乙酸（EDTA）中，大核染色质的热变性曲线在约76和83摄氏度处显示两个主要转变，类似于可溶性鸡红细胞染色质的解链。在相同离子强度条件下，将大核染色质的圆二色光谱与可溶性鸡红细胞染色质进行比较，发现非常相似。二维染色质/DNA琼脂糖凝胶电泳产生了一条DNA染色对角线，这在DNA和染色质电泳迁移率之间建立了严格的相关性。
