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G D Burkholder

Publications and source records attributed to G D Burkholder.

At least 37 records · Page 2Linked to original sources
PubMed1982

The effect of the chromosome banding techniques on the histone and nonhistone proteins of isolated chromatin.

The effect of a variety of chromosome banding techniques on the histone and nonhistone proteins of isolated, fixed, air-dried chromatin has been studied. Chromatin preparations were exposed to G-banding (SSC, urea, NaCl-urea, or trypsin), R-banding (Earle's BSS), and C-banding (NaOH or Ba(OH)2) treatments, and the proteins extracted from, and those remaining in the posttreatment chromatin, were examined by SDS polyacrylamide gel electrophoresis. The results indicate that: (i) each banding treatment produces specific quantitative and qualitative changes in the proteins of chromatin; (ii) the diverse treatments producing the same type of chromosome banding have both common and unique effects on the chromosomal proteins; and (iii) the treatments producing different types of chromosome banding have substantially different effects on the chromosomal proteins. It is not known whether these protein alterations are side effects occurring coincidentally with chromosome banding, or whether they are directly involved in mechanisms of banding. Results (ii) and (iii) suggest the latter possibility is more likely. If the protein alterations occur in specific chromosome regions, they may well influence the appearance of bands on mitotic chromosomes.

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PubMed1981

The ultrastructure of R-banded chromosomes.

Electron microscopy has been used to study the fine structural organization of R-banded chromosomes prepared by treatment of the chromosomes with a hot NaH2PO4 solution. The results indicate that there is a structural basis for R-banding with this technique. In comparison to untreated control chromosomes, the R-banded chromosomes had a greatly reduced electron density, suggesting that the heat treatment has a general adverse effect on chromosome structure. Chromatin fibers formed a coarse, irregular network throughout the chromosome and were often enlarged, probably as a result of the fusion of two or more native fibers. The chromatin fibers were more aggregated and had an increased electron density in the R-band regions of the chromosome than in the interbands. This indicates that the treatment has a differential effect on the structure of bands and interbands. A comparison of the ultrastructure of R- and G-banded chromosomes demonstrated that the distribution of aggregated chromatin was reversed by these two types of banding techniques; however, the treatments producing the R-banding appeared to induce less extreme differences in the degree of chromatin condensation in band and interband regions than those giving rise to G-banding. It is suggested that alterations of DNA-protein interactions may arise from the differential denaturation of proteins and/or DNA in R-band and interband regions during the heat pretreatment. Such differential alterations in DNA-protein interactions may induce localized changes in the organization of chromatin and may account for the subtle morphological differences observed between the band and interband regions.

Animals↗
PubMed1980

Proteins in chromosome banding. I. Effect of G-banding treatments on the proteins of isolated nuclei.

Nuclei were isolated from Chinese hamster cells, treated with hypotonic KCl, fixed in acetic methanol, and either air-dried in glass tubes (in situ) or left in suspension (in vitro). These preparations were then exposed to a variety of G-banding treatments, including the 2 xSSC, urea, NaCl-urea, and trypsin methods. The proteins extracted into the treatment solution and those remaining in the nuclei were analyzed by SDS polyacrylamide gel electrophoresis. The three former treatments extracted specific subsets of the total nuclear nonhistone proteins into the treatment solution. Some of the extracted nonhistones were common to all treatments while others were unique to a particular treatment. Variable amounts and types of the histones were also extracted by these treatments, but significant quantities of all of these proteins still remained in the nuclei afterwards. The trypsin treatment appeared to degrade some of the nonhistones, while other nonhistones, as well as the histones, were relatively resistant to trypsin digestion. Although there were a few differences in the residual proteins found in the nuclei after the various G-band treatments, the overall electrophoretic patterns of these proteins were generally similar. The results indicate that the G-banding techniques induce specific and reproducible changes in the proteins of isolated nuclei. If these banding treatments induce similar changes in the proteins of mitotic chromosomes, such alterations might be involved in mechanisms of chromosome banding.

Animals↗
PubMed1980

Proteins in chromosome banding. II. Effect of R- and C-banding treatments on the proteins of isolated nuclei.

SDS polyacrylamide gel electrophoresis was used to study the proteins extracted from, and those remaining in isolated, fixed, air-dried nuclei subjected to a variety of R- and C-banding techniques. The R-banding procedures, involving exposure to hot Earle's BSS or NaH2PO4, had the least effect of any of the banding techniques on the extraction of proteins from isolated nuclei. Only small amounts of 5 nonhistone proteins were detected in the Earle's BSS extract, and no proteins were found in the NaH2PO4 solution after treatment. The residual proteins remaining in the nuclei after either treatment were virtually identical to those in control nuclei. The C-banding techniques, on the other hand, produced substantial changes in the nuclear proteins. These techniques involve several sequential steps, including HCl treatment, exposure to NaOH or Ba(OH)2, and an incubation in hot SSC. The HCl treatment extracted a large variety of nonhistones and some of each of the remaining histones. No proteins were detected in the SSC solution. Some of the proteins extracted by Ba(OH)2 appeared after the two complete C-banding treatments revealed both similarities and differences. The Ba(OH)2 technique appeared to have a more severe effect on the nuclear proteins than the NaOH technique. Fewer residual nuclear proteins were observed after the former technique, but all of these were also represented in nuclei after the NaOH technique. The results indicate that the different treatments producing a common type of banding generally have similar effects on the nuclear proteins, while the treatments producing different types of banding (G-, R-, C-banding) have substantially different effects on these proteins. Such alterations may have implications for chromosome banding.

Animals↗
PubMed1980

Silver stained core-like structures in chinese hamster metaphase Chromosomes.

Chinese hamster metaphase chromosomes, subjected to prolonged hypotonic pretreatment and subsequently stained with ammoniacal silver, contained a darkly-stained core-like structure in each chromatid, surrounded by a halo of dispersed chromatin which was pale yellow to brown in color. The core was variable in its appearance, ranging from a continuous linear configuration to a spiral structure or a discontinuous, particulate structure. Within the centromeric regions, the cores frequently appeared more intensely stained than elsewhere in the chromosome. The nucleolus organizers also stained darkly and appeared to be attached to the core-like structures. It remains to be determined whether the cores represent a real component of metaphase chromosome structure, or whether they are artifacts resulting from abnormal chromatin aggregation arising at the time of chromosome preparation.

Animals↗
PubMed1977

Whole mount electron microscopy of the nucleolus in salivary gland cells of Drosophila melanogaster.

The nucleolus of Drosophila melanogaster salivary gland cells, examined by whole mount electron microscopy, consists of a fibrillar core region and a peripheral region containing both fibres and granules. These regions appear to correspond to the fibrillar and granular components, respectively, seen in thin sections. Most of the nucleoli were attached to the chromocenter region of the polytene chromosomes, containing the nucleolar organizer. Bundles of relatively straight chromatin fibres, 13 nm in diameter, extended from the chromocenter into the core region of the nucleolus, however it was not possible to trace the path of these chromatin fibres through the nucleolus since they were obscured within the mass of nucleolar fibres. The nucleolar fibres in both the core and peripheral regions were irregular and knobby, with a diameter of about 15 nm. In the core region, the fibres appeared to be of considerable length and were characteristically clustered together to form small interconnected masses. The fibres in the peripheral region were relatively short and some appeared to blend with amorphous, poorly-defined pools of material. Electron dense granules 15-20 nm in diameter were also associated with this amorphous substance. It is hypothesized that the formation and subsequent packaging of the 28s rRNA may be represented by a morphological transition of the peripheral fibres, via an amorphous pool-like intermediate stage, into the nucleolar granules. The results of this study indicate that whole mount electron microscopy may be a useful alternative to thin sectioning in high resolution studies of the nucleolus.

Animals↗
PubMed1976

Whole mount electron microscopy of polytene chromosomes from Drosophila melanogaster.

A simple method of whole mount electron microscopy has been developed to study the fine structural organization of Drosophila melanogaster polytene chromosomes. This method preserves the structural features of these chromosomes and may be of use for rapid gene localization at the ultrastructural level. Chromosome bands were electron dense regions composed of closely packed groups of chromomeres; thin bands consisted of a single row of transverse chromomeres, while thicker bands were composed of two to several rows of chromomeres. Interband regions contained relatively straight chromatin fibres which traversed the interband zone either singly or in bundles of several fibres. The interband chromatin fibres were generally 130 A in diameter or thicker, and appeared to be composed of two or more thinner (80-90 A) fibres. Presumptive puff regions were characterized by extended chromatin fibres having a longitudinal orientation, however some transverse rows of chromomeres were also seen in these regions, suggesting that not all of the chromomeres in a band may be involved in puff formation. The chromatin fibres inthe puffs were frequently thinner than those found in the interband regions. In stretched chromosomes, the chromatin fibres were drawn out into a mass of parallel fibres without any distinction between band and interband regions, supporting the hypothesis that individual chromatids are continuous through both band and interband regions and probably extend throughout the whole chromosome. No core fibres were observed in any of the polytene chromosomes studied.

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