Combined SYBR Green Real-Time with Telomeric Repeat Amplification Protocol (RQ-TRAP) to Detect Telomerase Activity
10.3969/j.issn.0253-9896.2014.08.003
- VernacularTitle:SYBR Green实时定量端粒重复序列扩增法检测端粒酶活性
- Author:
Wenqing MA
;
Fuzhi LIAN
;
Jinquan WANG
;
Lei YANG
- Publication Type:Journal Article
- Keywords:
telomere;
gene amplification;
enzyme-linked immunosorbent assay;
telomerase activity;
telomeric re-peat amplification protocol;
TRAP-ELISA
- From:
Tianjin Medical Journal
2014;(8):746-748
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish methodology to detect telomerase activity based on real-time quantitative PCR technique combined with telomeric repeat amplification protocol (TRAP). Methods RQ-TRAP system was developed by combining real-time quantitative PCR technique with conventional TRAP method. Telomerase activity was assessed and compared by RQ-TRAP assay and TRAP connected with enzyme-linked immunosorbent assay (TRAP-ELISA) respectively in 12 kinds of cells. Results The RQ-TRAP method was both accurate and specified in measuring telomerase activity in a series dilution of protein extracts from 293T cells. The sensitivity of this method was 8 cells and the amplification efficiency was 98.92%. Telomerase activity was not detected in negative control group. Statistical analysis revealed a strong correlation between the two assays (r2=0.762 5). Conclusion The feasibility of RQ-TRAP was proved in this article. Compared with TRAP-ELISA, RQ-TRAP has many advantages. Apart from sample extraction and real-time PCR cycling, no other extra time-consuming steps are needed for telomerase quantification;RQ-TRAP is less costly and more rapid and reliable than TRAP-ELISA for quantification of telomerase activity and it also support high throughput.
