Abstract 1234: Development of a quick sensitive real-time PCR assay for measuring telomere length

Telomeres are the ends of linear chromosomes with tandem hexameric nucleotide repeats. Its function is to protect the natural ends of chromosomes from being recognized as damaged DNA, hence contributing to chromosomal stability. Telomeric sequences shorten each time the DNA replicates; when the telomere shrinks to a certain threshold, the cell can no longer divide. Its metabolism slows down, the cell ages, and dies. This molecular clock mechanism is also associated with cancer progression and therapy. Turning off telomerase activity by shutting off telomerase activation in cancer cells will led to cellular senescence. The current gold standard method to measuring telomere length is by Southern blot; however, it is tedious and costly, and often takes up to two days to complete. Here, we describe a new, simplified automated real-time PCR assay to accurately detect the abundance of these 5’-TTAGGG-3’nucleotide repeats and determine frequency and the length of Telomere in 47 min. The assay demonstrated to be highly reproducible and accurate for human peripheral cells. The result has good correlation of R2=0.9939, slope=3.41, sensitivity to 1ng of gDNA and has an amplification efficiency of 100%. We evaluated this assay with 299 healthy human subjects to date ranging from 0 (core blood) to 99-year of age. The results are positive with an inverse correlation between telomere length and age, as telomeres were shorter with aging. The telomere length was measured in 13 samples using the gold standard Southern blot method and the new q-PCR method (R2 = 0.8623), significantly higher than the originally described (Cawthon NAS 2002). We believe this fast, sensitive methodology is a powerful tool and provides a new avenue for investigators to address the full extent and significance of telomere shortening in the tumorigenic process. This new assay described here will allow for more widespread use of this technique among cancer researchers and for study of age related telomere diseases and conditions. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1234. doi:10.1158/1538-7445.AM2011-1234