黑麦染色体的银染研究

在对黑麦染色体银染过程的盐酸解离条件进行探索的同时,对黑麦染色体的银染正反应区进行了研究,首次发现经短时间空气干燥(4-24h)的黑麦染色体制片,随着盐酸解离强度的递增,分别出现了核仁组织区(NOR)、NOR和端粒以及NOR和着丝点的银染正反应,就此现象讨论了端粒和着丝点的银染机理。     

黑麦染色体银染的初步研究

本文对黑麦染色体的银染条件进行了探索,并对黑麦染色体上银染正反应分布区进行了研究。首次发现黑麦染色体的核仁组成中心区、着丝点和端粒均能用硝酸银染色。对此现象的原因进行了讨论。     

松香草体细胞染色体银染色研究

松香草(Silphium perfoliatum, 2n=14)全部染色体着丝点和第5及第7对染色体短臂端部显示稳定的Ag-带。第1,2和6对染色体长臂居间区各有一居间带呈不稳定银染正反应。推测第5和第7对染色体为NOR染色体NOR分别位于两对染色体短臂末端。     

大麦染色体银带的研究

本文以六棱、四棱和二棱大麦为材料,用去壁低渗火焰干燥法制备染色体标本,标本在60℃恒温下经70—80％AgNO_3水溶液处理12—14小时,可在核仁组成区、着丝粒和端粒出现黑色银染区。细胞化学反应说明这些不同区域的银染物质具有相同的性质。银染带型与C带型和N带型迥然不同,3种大麦的银染带型也有差别。试验还证明染色体标本制备技术对银染效果影响极大,只有合适的酶解和火焰干燥处理才能促使着丝粒和端粒显示出银染正反应。     

蛙属中的一个特殊核型——双团棘胸蛙的核型及其C带和银带的研究

采用骨髓细胞直接制备染色体标本法和BSG、Ag-As显带技术。观察到双团棘胸蛙染色体数2n=64,NF=64,全部为端部着丝点染色体。银染显示的唯一1对核仁组织者(NOR)在第20号染色体的居间区,恰与该核型中出现的唯一1对次缢痕的位置一致。C带显现于几乎所有染色体的端部着丝点区,一部分核型在染色体的居间区和末端也出现C带。作者推测双团棘胸蛙很可能具有蛙属中至今所发现的最原始核型。     

白条草晰的染色体组型oC带和Ag-NOR。分析

白条草晰为2n二34T -I- 2m＋7.W,即常染色体中17对为端部着丝点大型染色体,I对为点状．性 染色体为zw 型。经BSG显带处理后,全部染色体的著丝点区都有职显的C带,同时还显示了10条 端粉C带,表明核型演化过程可能与染色体易位重组有关。银染结果仪显示1对NOR,它位于No. 17 染色体的末端。     

一种改进的植物染色体Ag—NOR染色方法及其应用

自1975年,Howell等和Goodpasture等报道人类染色体的Ag-NOR(银染核仁组成区)染色技术以来,银染色技术已广泛用于人类和哺乳动物染色体的核仁、核仁组成区、中心粒、着丝点、染色体轴心以及联会复合体等结构和行为的研究。但银染技术在植物     

BrdU和Hoechst 33258诱导黑麦染色体臂内银染区的研究

黑麦（Secale cereale)染色体除核仁组成区,着丝粒、端粒能被AgNt)3染色体,还可用BrdU和Hoechst 33258诱导出银染区,本文还对染色体差别银染的机制作了探讨。     

BrdU和Hoechst 33258诱导黑麦染色体臂内银染区的研究

黑麦（Secale cereale)染色体除核仁组成区,着丝粒、端粒能被AgNt)3染色体,还可用BrdU和Hoechst 33258诱导出银染区,本文还对染色体差别银染的机制作了探讨。     

人类双生子的银染核仁形成区的研究

近二十年来,人们为了探讨遗传与环境在 产生表型中的相互作用,已在解剖、生理、生化、 心理、行为及病理状态等各方面对人类双生子 进行了详尽的研究。1975年Goodpasture等⁵⁾ 应用银染技术使近端着丝粒染色体的副缴痕, 即人类核仁形成区（NOR）特异染色,并证卖 银染的位置是：RNA基因的位置。为了进一 步了解银染核仁形成区（Ag-NOR）和银染近 端着丝粒染色体联合（Ag-AA）的遗传特征, 我们用银染技术进行了双生子的Ag-NOR和 Ag-AA的研究。     

胃粘膜病变核仁组成区嗜银蛋白的银染计量和能谱分析

对77例胃粘膜活检标本进行银染,显示核仁组成区相关的嗜银蛋白(Ag-NOR),进行计数和统计分析,并对银染标本作电镜观察及能谱分析。结果表明,Ag-NOR 颗粒计数的多少,对胃肿瘤恶性程度的诊断有重要参考价值。电镜证实银染物质定位于核仁组成区,银染阳性区能谱分析显示 Ag 峰,说明银染反应具特异性.     

X-linked heterochromatin distribution in the holocentric chromosomes of the green apple aphid <Emphasis Type="Italic">Aphis pomi</Emphasis>

Chromatin organization in the holocentric chromosomes of the green apple aphid Aphis pomi has been investigated at a cytological level after C-banding, NOR, Giemsa, fluorochrome staining and fluorescent in situ hybridization (FISH). C-banding technique showed that heterochromatic bands are exclusively located on X chromosomes. This data represents a peculiar feature that clearly contradicts the equilocal distribution of heterochromatin typical of monocentric chromosomes. Moreover, silver staining and FISH carried out with a 28S rDNA probe localized rDNA genes on one telomere of each X chromosome; CMA₃ staining reveals that these silver positive telomeres are the only GC-rich regions among A. pomi heterochromatin, whereas all other C-positive bands are DAPI positive thus containing AT-rich DNA.     

Silver staining of the nucleolar organizer regions (NORs) on Lowicryl and cryo-ultrathin sections

Nucleolar organizer region (NOR) silver staining was applied to sections of fixed material. A positive reaction on cryo-ultrathin sections was found as well as on semithin and ultrathin Lowicryl sections. Repeatable staining that was easy to control was obtained by a one-step procedure after aldehyde-Carnoy fixation. Fixation of the material by formaldehyde and glutaraldehyde alone in cacodylate buffer also maintained reaction selectivity when ammonium chloride was used after fixation. Enzymatic digestion by pronase, RNase A, DNase I, or micrococcal nuclease was applied to ultrathin Lowicryl sections. Pronase digestion removed the silver-stained proteins, whereas digestion by the nucleases did not. A routine procedure is proposed for easy NOR silver staining of sections that preserves a good tissue ultrastructure and is also compatible with cytochemical and immunological investigations.     

Detection of fibrillarin in nucleolar remnants and the nucleolar matrix

In order to gain further insights into the fundamental structure of the nucleolus, nucleolar remnants of Xenopus and chickens were examined for the presence of fibrillarin and nucleolus organizer region (NOR) silver staining. Nucleolar remnants of Xenopus nucleated red blood cells were found to contain easily detectable amounts of fibrillarin and NOR silver staining. Upon examination of various tissues, fibrillarin and NOR silver staining were detected in nucleoli of Xenopus liver hepatocytes and within nucleoli of oocytes and follicle cells from ovaries of mature female toads. By comparison, nucleolar remnants of adult chicken nucleated red blood cells contained only trace amounts of fibrillarin and NOR silver staining, whereas red blood cell nucleolar remnants of immature chicks had easily detectable amounts of fibrillarin and NOR silver staining. Nucleoli from hepatocytes of both adult and immature chickens demonstrated comparable levels of fibrillarin and NOR silver staining. Since fibrillarin was found in nucleolar remnant structures, we tested for (and detected) its presence in residual nucleoli of in situ nuclear matrix derived from HeLa cells. These findings are discussed in terms of the basic structural and functional organization of the nucleolus.     

Silver staining in transcriptionally active NORs of meiotic and mitotic cells in Acheta domesticus (L.) (Orthoptera)

The behaviour of the NOR material in mitotic and meiotic cells of Acheta domesticus was studied by silver staining. — In mitotic chromosomes black silver staining is observed in the centromeric region of 2 pairs of acrocentric chromosomes. Additionally a polymorphic silver positive region is found at the telomere of a large submetacentric chromosome. — The Ag-pattern of the amplified rDNA material in various stages of oogenesis was followed. During pachytene the extra DNA body shows dark brownish staining and only a few black spots. One distinct black precipitate, however, is found in association with meiotic chromosomes. In early diplotene the central core of the extra DNA body is heavily stained with silver. The outer shell shows only brown staining. In the following stages of diplotene the compact structure of the outer shell is loosened and small brown extra nucleoli are found in the remaining nucleus. These nucleoli show black Ag-precipitates in their centres. During the desintegration of the extra DNA body the nucleus becomes filled with small extra nucleoli. The black stained central core is reduced in size and finally disappears.     

The Effect of Ph on Silver Staining of Nucleolus Organizer Regions

The effect of pH on silver staining of the nucleolus organizer regions (NORs) of human chromosomes has been investigated between pH 6.5 and 12.0. Nonvolatile mixtures of ethanolamine and ethanolammonium nitrate replaced the ammonia of standard procedures. The optimal NOR staining obtained at pH 3.5 by the silver staining procedure of Howell and Black served as a standard; this procedure stained all ten NORs in 90% of mitoses. Similar NOR staining was found in 75% of mitoses stained at pH 11.7 or 11.8, but only in 10-15% of mitoses stained between pH 11.6 and 10.0. Between pH 10.0 and 9.0 NOR staining was incomplete, and between pH 8.5 and 6.5 there was no NOR staining.     

The effect of pH on silver staining of nucleolus organizer regions

The effect of pH on silver staining of the nucleolus organizer regions (NORs) of human chromosomes has been investigated between pH 6.5 and 12.0. Nonvolatile mixtures of ethanolamine and ethanolammonium nitrate replaced the ammonia of standard procedures. The optimal NOR staining obtained at pH 3.5 by the silver staining procedure of Howell and Black served as a standard; this procedure stained all ten NORs in 90% of mitoses. Similar NOR staining was found in 75% of mitoses stained at pH 11.7 or 11.8, but only in 10-15% of mitoses stained between pH 11.6 and 10.0. Between pH 10.0 and 9.0 NOR staining was incomplete, and between pH 8.5 and 6.5 there was no NOR staining.     

Nucleolar activity in differentiated cells after stimulation

Initiation of nucleolar organizer region (NOR) activity was observed by using the silver staining method at various times after activation or stimulation of differentiated cells. Two methods were used: (1) activation of human lymphocytes by treatment with phytohemagglutinin (PHA), and (2) cell-cell fusion of chick erythrocytes with squirrel monkey cells. An increase in NOR activity in lymphocytes was seen as early as 4 h after PHA treatment and between 10 and 22 h in the chick erythrocytes after fusion. In both systems, as the size of the dormant cell nucleus increased, the amount of silver staining increased until the silver-stained area approached that of cycling cells.     

Procedures for specific detection of silver-stained nucleolar proteins on western blots.

Nucleolar organizer region (NOR)-silver staining of the chromosomes and nucleoli is a method that enables the detection of proteins associated with the ribosomal genes. We adapted the most commonly used cytochemical NOR-silver staining techniques to Western-blotted proteins of HeLa cells, mimicking the silver staining of cells in situ, and testing several parameters that may influence the in situ reaction. Two of these techniques, both one-step methods with colloidal developers, were standardized to obtain reproducible results. The specificity of NOR staining is documented by: (a) only a few bands are revealed among the many proteins detected by total proteins staining on gels or blots; two major groups of bands are found around 100 KD and 40 KD that could correspond at least in part to nucleolin and B23 nucleolar proteins; (b) the silver staining of bands was not the result of the high relative protein concentrations; and (c) the same number of NOR-silver-stained bands was observed across a large range of protein concentrations. The reaction appeared to be specific for a subset of nucleolar proteins, because the same bands were observed with the use of nucleolar, nuclear, or total cell protein extracts, and the silver grains observed in electron microscopy were clearly confined to the nucleolar fibrillar centers and dense fibrillar component. The efficiency of the reaction was not modified by any of the tested fixative pre-treatments except that involving methanol. The presented standardization of NOR-silver staining on Western blots allows the characterization of the Ag-NOR proteins and their specific regions responsible for silver staining of the nucleolus.