Estimation of copy number using SYBR Green: confounding by AT-rich DNA and by variation in amplicon length
Colborn, J.M.; Byrd, B.D.; Koita, O.A.; Krogstad, D.J.
American Journal of Tropical Medicine and Hygiene 79(6): 887-892
2008
ISSN/ISBN: 0002-9637 PMID: 19052298 Document Number: 621104
Although SYBR Green is used to estimate copy number, its fluorescence varies with amplicon length and adenine/thymine (AT) content. As a result, threshold cycle (Ct) values obtained using real-time polymerase chain reaction (PCR) are lower for longer amplicons (P<0.001) and amplicons with greater AT content (P<0.001). In contrast, neither amplicon length nor AT content affects the Ct with TaqMan probes or LUX-labeled primers.