Christopher J. Bostock

Specific arrangements of human satellite III DNA sequences in human chromosomes.

DNA was extracted from various rodent-human somatic cell hybrids that contained single or a few human chromosomes. These DNAs were examined by a combination of restriction endonuclease digestion, gel electrophoresis, and filter hybridisation to radioactive satellite DNA probes following transfer of the denatured restriction fragments from a gel to a nitrocellulose filter. In this way the arrangement of sequences homologous to human satellite III were examined on human chromosomes 1, 7, 11, 15, 22 and X. It was found that the distribution of restriction endonuclease sites within satellite III DNA is different on different chromosomes.

A study of sequence homologies in four satellite DNAs of man.

Abstract Satellites I, II, III and IV (Corneo et al. , 1968,1970,1971) have been purified from human male placental DNA. The sequences present in these four DNA components have been characterized by analytical buoyant density, thermal denaturation, DNA reassociation, DNA hybridization and gel electrophoresis coupled with hybridization following either Hae III or Eco RI restriction endonuclease digestion. Satellites III and IV were found to be virtually indistinguishable by a variety of criteria. Cross-satellite reassociation showed that 40% of the molecules present in satellite III contain sequences that are homologous to 10% of the molecules of either satellite I or satellite II. Reassociated satellite I melts as a single component, as do the hybrid duplexes between satellite I and satellite III. In contrast, reassociated satellites II, III and IV, and the hybrid duplexes formed between satellites II and III and between satellites II and IV, melt as two distinct components with different thermal stabilities. Digestion of satellite III with Hae III gives rise to a series of fragments whose sizes are 2, 3, 4, 5, 6, 7, 8 and 11 times the size of the smallest 0.17 × 10 3 basepair fragment, in addition to a 3.4 × 10 3 base-pair male-specific fragment (Cooke, 1976) and high molecular weight material. The sequences contained in the fragments of the Hae III ladder are diverged from each other as well as being non-homologous with those of the 3.4 × 10 3 base-pair and high molecular weight fragments. The latter contain Eco RI recognition sites. Satellite II has a similar pattern of fragments to satellite III following digestion with Hae III, although it can be distinguished from satellite III on the basis of the products of Eco RI digestion. Satellite I contains neither Hae III nor Eco RI recognition sites. The cross-satellite homologies of the sequences present in fragments of differing sizes produced by restriction enzyme digestion have also been studied.

Analysis of the frequency of sister chromatid exchange in different regions of chromosomes of the kangaroo rat (Dipodomys ordii).

The frequency of sister chromatid exchanges (SCEs) has been determined for C band and non-C band regions of chromosomes of the kangaroo rat after staining with the fluorescence plus giemsa (FPG) technique. After one complete round of DNA synthesis in the presence of bromodeoxyuridine (BrdU) staining of the C band regions revealed simple or complex asymmetries between chromatids. After two complete rounds of DNA synthesis in the presence of BrdU “harlequin” chromosomes were observed. Analysis of the distribution of SCE in chromosomes at their 1st and 2nd mitosis showed that relatively few exchanges occur within C band regions, although the frequency of SCEs is high at the junction between C band and non-C band chromosome regions.

Gene amplification in methotrexate-resistant mouse cells: II. Rearrangement and amplification of non-dihydrofolate reductase gene sequences accompany chromosomal changes

Abstract Total amplified DNA in methotrexate-resistant mouse lymphoma EL4 cells and mouse melanoma PG19 cells has been characterized in two ways. Metaphase spreads show the presence of additional chromosome forms that are either “homogeneously staining” chromosomes or “double minute” and ring chromosomes. Gel electrophoresis of restriction enzyme-digested nuclear DNA shows the presence of amplified sequences, the pattern of which is unique in each of five cell lines. We conclude that extensive DNA rearrangement has taken place during amplification.

Gene amplification in methotrexate-resistant mouse cells: III. Interrelationships between chromosome changes and DNA sequence amplification or loss

Abstract The amplified DNA sequences within a single subline of methotrexate-resistant mouse cells have a complex intracellular location, some being associated with “double minute” chromosomes, and some with normal-sized chromosomes. However, within a cell line and probably within a cell their internal structure is stable and homogeneous. These results, and those from the previous paper (Bostock & Tyler-Smith, 1981), suggest that gene amplification in these cells has two distinct stages. Firstly, an amplifiable unit is generated by a process involving DNA rearrangement. Secondly, the unit increases in number but remains unchanged in structure.

S phase patterns of replication of different satellite DNAs in three species of Dipodomys (kangaroo rat)

The pattern of synthesis of four different DNA components, main band, MS, HS-α and HS-β satellites has been studied in each of three different species of kangaroo rat: Dipodomys ordii, Dipodomys panamintinus and Dipodomys merriami . All DNA components are synthesized to some extent at all S phase intervals, but they all show rapid changes in the rate of synthesis at defined points in the S phase. Within one species, different satellite DNAs are replicated at their maximal rate during different intervals of the S phase and are not always late replicating. The time of synthesis of HS-α satellite DNAs is independent of species type, but HS-β satellite DNA is synthesized at the maximal rate during a different interval of the S phase in each of the three species.