Walmsley M E, Davies H G
J Cell Sci. 1975 Jan;17(1):113-39. doi: 10.1242/jcs.17.1.113.
Adult hen erythrocyte nuclei are isolated from cells or haemolysed in situ by acting on the plasma membrane with rotating knives or with non-ionic detergents. When the isolation medium contains magnesium ions (1 mM), sucrose (0-4 M) and Tris buffer (0.01 M, pH 7-5) called SMTOG (see text), the ultrastructure in thin sections through the condensed chromatin bodies, after staining with either uranyl-lead or phosphotungstic acid (PTA), is similar to that found in the intact cell. Hence it can be concluded that the 2 phases which comprise chromatin, the o- and e-phase, survive nuclear isolation. These are so called because the structural units in chromatin are arranged at the surface of the nucleus into one or more layers and give rise to oddly (o) and evenly (e) numbered bands. The 0-phase is also largely retained after extensive washing in 0-07 M NaC1 as shown by electron microscopy and biochemical measurements; only 6% of the total nuclear protein is removed, a value small compared with the fractional amount of the chromatin protein calculated to lie in the o-phase, about 70%. After extensive washing in saline-EDTA there are structural changes in chromatin, but biochemical data show that the molecules in the o-phase are also largely retained; loss of protein amounts to between 5 and 11%. These data suggest that the o-phase is a structural component of the chromatin bodies. They support the hypothesis that condensed chromatin is formed by folding superunit threads. These units consist of a central thread-like element about 17 nm diameter which stains preferentially with uranyl-lead and forms the e-phase, with an outer cylindrical shell forming the o-phase of total diameter about 28nm. The 5-10% proteins removed by salt washes are located exclusively in a particulate component, quite likely the chromatin. They have been examined by sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis. There are about 10 or more protein species, ranging in molecular weight from 21000 upwards. The groups of large granules previously found in the nuclear sap of intact erythrocytes are shown to be associated with an amorphous or finely fibrillar body.
成年母鸡红细胞核可通过用旋转刀作用于质膜或用非离子去污剂原位溶血从细胞中分离出来。当分离介质含有镁离子（1 mM）、蔗糖（0 - 4 M）和Tris缓冲液（0.01 M，pH 7.5），即所谓的SMTOG（见正文）时，在用铀铅或磷钨酸（PTA）染色后，通过浓缩染色质体的薄切片中的超微结构与完整细胞中的相似。因此可以得出结论，构成染色质的两个阶段，即o期和e期，在核分离后仍然存在。之所以这样称呼，是因为染色质中的结构单元在核表面排列成一层或多层，并产生奇数（o）和偶数（e）编号的带。如电子显微镜和生化测量所示，在0.07 M NaCl中广泛洗涤后，o期也基本保留；仅去除了总核蛋白的6%，与计算得出的位于o期的染色质蛋白的分数相比，这个值很小，约为70%。在盐水 - EDTA中广泛洗涤后，染色质有结构变化，但生化数据表明o期的分子也基本保留；蛋白质损失在5%至11%之间。这些数据表明o期是染色质体的结构成分。它们支持这样的假设，即浓缩染色质是由超单位细丝折叠形成的。这些单位由一个直径约17 nm的中心丝状元件组成，该元件优先用铀铅染色并形成e期，还有一个外部圆柱形外壳形成总直径约28 nm的o期。通过盐洗去除的5% - 10%的蛋白质仅位于颗粒成分中，很可能是染色质。它们已通过十二烷基硫酸钠（SDS）聚丙烯酰胺凝胶电泳进行了检测。有大约10种或更多的蛋白质种类，分子量从21000以上不等。先前在完整红细胞核液中发现的大颗粒群被证明与无定形或细纤维状物体有关。
