Critical evaluation of HPV16 gene copy number quantification by SYBR green PCR.

ABSTRACT:

Background

Human papilloma virus (HPV) load and physical status are considered useful parameters for clinical evaluation of cervical squamous cell neoplasia. However, the errors implicit in HPV gene quantification by PCR are not well documented. We have undertaken the first rigorous evaluation of the errors that can be expected when using SYBR green qPCR for quantification of HPV type 16 gene copy numbers. We assessed a modified method, in which external calibration curves were generated from a single construct containing HPV16 E2, HPV16 E6 and the host gene hydroxymethylbilane synthase in a 1:1:1 ratio.

Results

When testing dilutions of mixed HPV/host DNA in replicate runs, we observed errors in quantifying E2 and E6 amplicons of 5-40%, with greatest error at the lowest DNA

SUBMITTER: Roberts I 

PROVIDER: S-EPMC2529285 | biostudies-literature | 2008 Jul

REPOSITORIES: biostudies-literature