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1.
We have established a series of 20 colorectal cancer cell lines and performed cytogenetic and RFLP analyses to show that the
recurrent genetic abnormalities of chromosomes 1, 5, 17 and 18 associated with multistep tumorigenesis in colorectal cancer,
and frequently detected as recurrent abnormalities in primary tumours, are also retained in long-term established cell lines.
Earlier studies by us and other investigators showed that allelic losses of chromosomes 1 and 17 in primary colorectal cancers
predicted poorer survival for the patients (P = 0.03). We utilized the cell lines to identify specific chromosomal sites or gene(s) on chromosomes 1 and 17 which confer
more aggressive phenotype. Cytogenetic deletions of chromosome 1p were detected in 14 out of the 20 (70%) cell lines, whereas
allelic deletions for 1p using polymorphic markers were detected in 13 out of 18 (72%) informative cell lines for at least
one polymorphic marker. We have performed Northern blotting, immunohistochemical staining (p53 mRNA, protein) and RFLP analysis
using several probes including p53 and nm23. RFLP analysis using a total of seven polymorphic markers located on 17p and 17q
arms showed allelic losses aroundthe p53 locus in 16 out of the 20 cell lines (80%), four of which were losses of thep53 locus itself. In addition, seven cell lines (out of nine informative cases) also showed losses of thenm23 gene, four with concurrent losses of thep53 locus, while the remaining three were homozygous. In addition, five out of seven cell lines withnm23 deletions were derived from hepatic metastatic tumours, and one cell line was obtained from recurrent tumour. A comparison
between allelic deletions of 1p and functional loss ofnm23 gene revealed a close association between these two events in cell lines derived from hepatic metastasis. Following immunohistochemical
staining, nine out of the twenty cell lines showed high levels (25–80%) of mutant p53, four showed intermediate levels (>20%),
and seven had undetectable levels of the protein. Of these seven, four showed complete absence of mRNA. Of the remaining three
cell lines one showed aberrant mRNA due to germline rearrangement of thep53 gene, whereas in two cell lines normal levels of mRNA were present. Nineteen of the 20 cell lines had normal germline configurations
for thep53 gene, while one showed a rearrangement. These data suggest that functional loss ofp53 andnm23 genes accomplished by a variety of mechanisms may be associated with poor prognosis and survival. In addition, concurrent
deletions of chromosome regions 17p, 17q and 1p were closely associated with high-stage hepatic metastatic disease. These
cell lines with well-characterized genetic alterations and known clinical history provide an invaluable source of material
for various biological and clinical studies relating to multistep colorectal tumorigenesis. 相似文献
2.
Appropriate final disposal of sewage sludge (SS) generated by wastewater treatment plants (WWTP) has been considered a serious environmental problem, but also a viable alternative to be applied in agriculture, once SS is rich in organic matter and nutrients. However, SS can be a source of contamination of several toxic agents. Therefore, its use in agriculture requires special care to avoid possible damage to the environment and exposed organisms. Detoxification of toxic wastes can be performed using the monitored natural attenuation, which involves biological, physical and chemical processes that frequently occur in the environment. This study aimed to assess the feasibility of decontaminating SS after different periods of monitored natural attenuation. To this end, samples of SS and associations of soil/SS with proportions of 10, 25 and 50% SS were buried for 0, 2, 6 and 12 months in holes prepared in a place free of contamination. Allium cepa was used as an indicator to assess the efficiency of the natural attenuation process. According to chemical analysis, the SS samples presented a high concentration of m- and p-cresol, especially for samples analyzed after 0 or 2 months of natural attenuation. The microorganisms present in the SS belonged to 17 different genera of bacteria, which varied in the microbial composition among samples. Both, raw SS and aqueous SS extracts induced DNA damage in A. cepa, even when associated with soil. However, this effect was observed to decline during the attenuation period, although significant effects were detected for the highest tested concentration (100% SS) even at the end of this process. These results thus indicated the necessity of applying a stabilization process associating SS and soil for a period of at least 12 months and showed that the studied raw SS is not a viable material for use as a soil reconditioner, even after natural attenuation. A. cepa test proved to be a useful tool to assess the efficiency of SS detoxification process. Therefore, we suggest that the application of SS in agriculture should be approached with caution and that the SS must be previously submitted to methodologies that evaluate its toxic potential. 相似文献
3.
Abstract Broad host-range RK2-based cosmid vectors ('costramids') are increasingly used in molecular genetic studies of Gram-negative soil bacteria such as Rhizobium spp. we describe a simple modification of existing methods, whereby a genomic library constructed in a stringently replicated vector can be screened for genes which are undetectable by colony hybridization due to background cross-hybridization. This method allows the use of 'heterologous' probes (interspecies hybridization) to isolate several presumptive genes of interest from a gene bank of Rhizobium sp. NGR234 made in the costramid pRK7813. These are a gene with homology to the citrate synthase gene ( gltA ) or Escherichia coli , the gene encoding δ-aminol evulinic acid synthase ( hemA ), and a gene or genes regulating dicarboxylate transport. 相似文献
4.
5.
D. L. Zhang K. Q. Li W. Gu L. F. Hao 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1987,75(1):132-137
Summary Different phytohormone concentrations induced different fequencies of various chromosome aberrations in calli of Vicia faba. NAA 10 ppm plus KT 2.5 ppm produced more haploids and NAA 30 ppm plus NAA 7.5 ppm produced more tetraploids and breakage. The relationship among the aberrations was analyzed. The hypothesis of ploidy equilibrium was established. The chromosome doubling rate and reduction rate of each treated group were calculated in relation to the observed data and the hypothesis. The frequency of tetraploids and breakage are correlated with each other. The frequency of total aberrations is linearly correlated with that of micronucleus formation. The regression equation is x=31.92+ 10.67 y. 相似文献
6.
Herbert N. Arst Jr. David Tollervey Mark X. Caddick 《Molecular & general genetics : MGG》1989,215(2):364-367
Summary The areA
r
-18 mutation is a loss-of-function mutation in areA, the positive acting regulatory gene mediating nitrogen metabolite repression in Aspergillus nidulans. It results from a reciprocal translocation which splits the coding region into 5 and 3 moieties. Surprisingly, we have selected rare intracistronic revertants of areA
r
-18. From crosses heterozygous for areA
r
-18 revertant alleles, duplication-deficiency progeny containing two copies of a substantial portion of chromosome IV but lacking part of chromosome III, including the 5 moiety of areA, have been obtained. For all four revertants analysed genetically, growth properties of these duplication-deficiency strains indicate that the reversion events involve the 3 portion of areA and that the 5 portion of areA is unnecessary for the revertant phenotype. This conclusion was directly confirmed for one revertant using Southern blotting. As all four reversion events involve additional chromosomal rearrangements, they probably fuse functional promoters, ribosome binding sites and in frame initiation codons to the 3 portion of the gene. In the course of characterisation of these mutations, new mapping data for a large region of chromosome IV have been generated, and a new reciprocal translocation activating the cryptic regulatory gene areB, whose product can substitute for that of areA, has been identified. 相似文献
7.
Philippe Lejeune Philippe Bertin Corinne Walon Karine Willemot Charles Colson Antoine Danchin 《Molecular & general genetics : MGG》1989,218(2):361-363
Summary Spontaneous mutants of Escherichia coli K12 displaying an increased level of the kanamycin resistance conferred by plasmid pGR71 were selected. Several mutants obtained in this way apparently carry large chromosomal deletions extending into galU and/or bglY (27 min). This positive selection of deletions allowed detection of a new locus located between galU and bglY. Deletions of this locus are responsible for increased resistance to kanamycin (Irk), decreased resistance to l-serine in minimal medium (Drs) and decreased resistance to chloramphenicol (Drc) when a cat gene is present in the bacteria. 相似文献
8.
Francis Fabre Nieve Magana-Schwencke Roland Chanet 《Molecular & general genetics : MGG》1989,215(3):425-430
Summary The RAD18 gene of Saccharomyces cerevisiae is involved in mutagenic DNA repair. We describe its isolation from a yeast library introduced into the centromeric YCp50 vector, a low copy number plasmid. The insert was sublconed into YCp50 and into the multicopy YRp7 plasmid. RAD18 is not toxic when present in multiple copies but the UV survival response indicates an heterogeneity in the cell population, a fraction of it being more sensitive. A DNA segment, close to RAD18, is toxic on the multicopy plasmid and may correspond to the tRAN sup61 known to be tightly linked to RAD18. Chromosomal deletions of RAD18 were constructed. The gene is not essential and the deleted strains have the properties of single site mutants. Thus, RAD18 appears to be essentially involved in DNA repair metabolism. 相似文献
9.
Atsuko Matsuoka Akiko Hirosawa Shinasku Natori Shigeo Iwasaki Toshio Sofuni Motoi Ishidate Jr. 《Mutation research》1989,215(2):179-185
The chromosomal aberration test using a Chinese hamster lung cell line (CHL) was carried out on ptaquiloside and its related compounds, hypoloside B, hypoloside C, illudin M and illudin S. Ptaquiloside induced chromosomal aberrations at doses as low as 4.5 μg/ml (0.0113 mM). The clastogenic effect was ph-dependent. The same activity was observed at a 90-fold higher dose at pH 5.3 in the culture medium compared with the activity at pH 74. or pH 8.0. Both hypoloside B and hypoloside C were also clastogenic at almost the same dose levels as that of ptaquiloside. Illudin M and illudin S were also potet clastogens and induced aberrations at much lower doses than ptaquiloside. These results suggest that the clastogenic effect is involved in the mechanism of carcinogenic potency of ptaquiloside in animals. 相似文献
10.
Chromosomal nonhistone high-mobility-group (HMG) proteins were purified from nuclei of maize (Zea mays L. cv. A619) endosperm and leaf tissue. Tissuespecific differences were observed in their polypeptide patterns, in in-vitro phosphorylation experiments with a casein-kinase type II, and by Western blot analysis with antisera against different HMG proteins. Gelfiltration chromatography demonstrated that maize HMG proteins occur as monomers. By measuring the capacity of the HMG proteins to bind to the 5 flanking region of a zein gene, the sensitivity of the proteins to different temperatures, salt concentrations and pH values was determined.Abbreviations EMSA
electrophoretic-mobility-shift assay
- FPLC
fast protein liquid chromatography
- HMG
high-mobility group
- kDa
kilodaltons
- PVDF
polyvinylidenedifluoride
- SDS-PAGE
sodium dodecyl sulfate-polyacrylamide gel electrophoresis
We would like to thank Mrs. E. Brutzer for excellent technical assistance. We are indebted to Mrs. M. Strecker and Dr. W. Bessler of the Institut für Immunbiologie, Freiburg, FRG, for the preparation of antisera and we gratefully acknowledge helpful discussions with Drs. T. Quayle, R. Grimm and U. Müller of this institute. This work was supported by grants from the Deutsche Forschungsgemeinschaft and the Fond der Chemischen Industrie. 相似文献