A procedure is described which gives clean chromatin preparations from the free-living nematode Caenorhabditis elegans. It involves homogenization using glass beads, collection of the precipitate from a low speed centrifugation, removal of cell membranes with Triton X-100, several washes with 0.14 M NaCl, sucrose density gradient centrifugation, a cycle of extraction and reprecipitation using dilute Tris buffer and 0.14 M NaCl respectively, and final extraction of the purified deoxyribonucleoprotein in 10 mM Tris-HCl (pH 8). 2. Acidic urea gel electrophoresis of the histones from C. elegans yielded 4 main groups which were preliminary identified as H1, H2a (+ H3?), H2b, H4 and moved on the gels in that order of increasing mobility. the coincidence of histone H3 with H2a was putative, but its presence was firmly suggested by the generation of a dimeric form in oxidizing conditions. 3. By SDS-Tris-glycine gel electrophoresis of the non-histone chromosomal proteins of C. elegans, about 18 proteins were distinguished with molecular weights ranging from 15,000 to 100,000 daltons.
摘要
本文描述了一种从自由生活的线虫秀丽隐杆线虫中获取纯净染色质制剂的方法。该方法包括使用玻璃珠进行匀浆,低速离心收集沉淀,用Triton X - 100去除细胞膜,用0.14 M NaCl洗涤数次,蔗糖密度梯度离心,分别使用稀Tris缓冲液和0.14 M NaCl进行提取和再沉淀循环,以及最终在10 mM Tris - HCl(pH 8)中提取纯化的脱氧核糖核蛋白。2. 对秀丽隐杆线虫的组蛋白进行酸性尿素凝胶电泳,产生了4个主要组,初步鉴定为H1、H2a(+ H3?)、H2b、H4,并在凝胶上以迁移率增加的顺序移动。组蛋白H3与H2a的重合是推测性的,但在氧化条件下产生二聚体形式有力地表明了它的存在。3. 通过对秀丽隐杆线虫的非组蛋白染色体蛋白进行SDS - Tris - 甘氨酸凝胶电泳,区分出了约18种蛋白质,其分子量范围为15,000至100,000道尔顿。