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J Korean Soc Ther Radiol Oncol > Volume 23(1); 2005 > Article
The Journal of the Korean Society for Therapeutic Radiology and Oncology 2005;23(1): 43-50.
Identification of Differentially Expressed Radiation-induced Genes in Cervix Carcinoma Cells Using Suppression Subtractive Hybridization
Jun Sang Kim, Young Sook Lee, Jeung Hoon Lee, Woong Hee Lee, Eun Young Seo, Moon June Cho
1Departments of Radiation Oncology, Chungnam National University, Daejeon, Korea. mjcho@cnu.ac.kr
2Departments of Dermatology, College of Medicine, Chungnam National University, Daejeon, Korea.
3Departments of Biology, College of Natural Sciences, Chungnam National University, Daejeon, Korea.
4Cancer Research Institute, College of Medicine, Chungnam National University, Daejeon, Korea.
A number of genes and their products are induced early or late following exposure of cells to ionizing radiation. These radiation-induced genes have various effects on irradiated cells and tissues. Suppression subtractive hybridization (SSH) based on PCR was used to identify the differentially expressed genes by radiation in cervix carcinoma cells. MATERIASL AND
Total RNA and poly (A)+ mRNA were isolated from irradiated and non-irradiated HeLa cells. Forward- and reverse-subtracted cDNA libraries were constructed using SSH. Eighty-eight clones of each were used to randomly select differentially expressed genes using reverse Northern blotting (dot blot analysis). Northern blotting was used to verify the screened genes.
Of the 176 clones, 10 genes in the forward-subtracted library and 9 genes in the reverse-subtracted library were identified as differentially expressed radiation-induced genes by PCR-select differential screening. Three clones from the forward-subtracted library were confirmed by Northern blotting, and showed increased expression in a dose-dependent manner, including a telomerase catalytic subunit and sodium channel-like protein gene, and an ESTs (expressed sequence tags) gene.
We identified differentially expressed radiation-induced genes with low-abundance genes with SSH, but further characterization of theses genes are necessary to clarify the biological functions of them.
Key Words: Cervix carcinoma, Suppression subtractive hybridization, HeLa cells, Radiation-induced genes
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