Evaluation of Commercial Complementary DNA Synthesis Kits for Detecting Human Papillomavirus
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초록
Cervical cancer is the fourth most common malignant neoplasm in women worldwide. Most cases of cervical cancer are caused by an infection by the human papillomavirus. Molecular diagnostic methods have emerged to detect the HPV for sensitivity, specificity, and objectivity. In particular, real-time PCR has been introduced to acquire a more sensitive target DNA or RNA. RNA extraction and complementary DNA synthesis are proceeded before performing real-time PCR targeting RNA. To identify an adequate and sensitive cDNA synthesis kit, this study evaluated the two commonly used kits for cDNA synthesis. The results show that the R2 and efficiency (%) of the two cDNA synthesis kits were similar in the cervical cancer cell lines. On the other hand, the Takara kit compared to Invitrogen kit showed P＜0.001 in the 102 and 103 SiHa cell count. The Takara kit compared to the Invitrogen kit showed P＜0.001 in the 101 and 102 HeLa cell count. Furthermore, 8, 4, 2, 1, and 0.5 ml of forty exfoliated cell samples were used to compare the cDNA synthesis kits. The Takara kit compared to the Invitrogen kit showed P＜0.01 in 8, 4, and 1 ml and P＜0.05 in 0.5 mL. The study was performed to identify the most appropriate cDNA synthesis kit and suggests that a cDNA synthesis kit could affect the real-time PCR results.
Abstract
Cervical cancer is the fourth most common malignant neoplasm in women worldwide. Most cases of cervical cancer are caused by an infection by the human papillomavirus. Molecular diagnostic methods have emerged to detect the HPV for sensitivity, specificity, and objectivity. In particular, real-time PCR has been introduced to acquire a more sensitive target DNA or RNA. RNA extraction and complementary DNA synthesis are proceeded before performing real-time PCR targeting RNA. To identify an adequate and sensitive cDNA synthesis kit, this study evaluated the two commonly used kits for cDNA synthesis. The results show that the R2 and efficiency (%) of the two cDNA synthesis kits were similar in the cervical cancer cell lines. On the other hand, the Takara kit compared to Invitrogen kit showed P＜0.001 in the 102 and 103 SiHa cell count. The Takara kit compared to the Invitrogen kit showed P＜0.001 in the 101 and 102 HeLa cell count. Furthermore, 8, 4, 2, 1, and 0.5 ml of forty exfoliated cell samples were used to compare the cDNA synthesis kits. The Takara kit compared to the Invitrogen kit showed P＜0.01 in 8, 4, and 1 ml and P＜0.05 in 0.5 mL. The study was performed to identify the most appropriate cDNA synthesis kit and suggests that a cDNA synthesis kit could affect the real-time PCR results.
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