Expression of exon 13 from the Ki-67 gene in human cells and tissues by digoxigenin-labelled mRNA in situ hybridization
- Author:
- No author available
- Journal Title:
- CHINESE MEDICAL JOURNAL
- Issue:
- 1
- DOI:
- 10.3760/cma.j.issn.0366-6999.2001.01.108
- Key Word:
- Ki-67基因;mRNA原位杂交;MIB-1;细胞增殖;in situ hybridization;Ki-67;mRNA;MIB-1;cell proliferation
Abstract： Objective To get insight on the regulatory mechanism of Ki-67 gene expression in malignant cell cycle. Methods Non-radioactive in situ hybridization (ISH) was undertaken, combined with immunohistochemistry to study the Ki-67 gene transcription and translation in various human cells and tissues. HeLa cells and fresh colon cancer cells, tonsil, normal pancreas and pancreatic cancer tissues were used in this study. A 435?bp cDNA fragment located in exon 13 of the Ki-67 antigen gene was amplified by polymerase chain reaction (PCR). Digoxigenin-labelled antisense and sense RNA probes were prepared for detecting Ki-67 mRNA, combined with MIB-1 immuno～histochemistry.Results Successful localization of Ki-67 mRNA in human HeLa cells, colon cancer cells, tissues specimen of the tonsil and pancreatic cancer tissue sections was accomplished by digoxigenin-labelling in situ hybridization technique. ISH to colon cancer cells and pancreatic cancer tissue slides showed that much stronger cytoplasm and perinuclear mRNA signals of the Ki-67 gene were present in malignant cells than in normal cells, which was in accordance with MIB-1 nuclear protein signals.Conclusions A sensitive and practical in situ hybridization method for the analysis of Ki-67 antigen mRNA in human cell and tissue was developed. Abnormal transcription of exon 13 of Ki-67 gene might be responsible for malignant cell proliferation in colon and pancreatic cancer.
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