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Revista Cubana de Medicina Tropical

Print version ISSN 0375-0760On-line version ISSN 1561-3054

Abstract

MARTINEZ RODRIGUEZ, Pedro Ariel et al. Standardization of a real-time polymerase chain reaction system for quantification of human herpesvirus 8. Rev Cubana Med Trop [online]. 2009, vol.61, n.2. ISSN 0375-0760.

OBJECTIVE: to standardize a real-time polymerase chain reaction system to determine the human herpes virus 8 viral load in several samples from patients suspected of this type of infection. METHODS: three internationally known methods were evaluated to obtain standard DNA in standard external curve constructions, which allow determining the number of target DNA copies in the suspected samples. RESULTS: three standards DNA were obtained from cloning ORF26 gene fragment of human herpesvirus 8 in a vector (plasmid DNA), with the use of purified polymerase chain reaction products and of genomic DNA of BCBL cell lines. The pattern curves were constructed on the basis of each of the resulting standard DNA, which showed strong linear correlation (r= -1) and very low error values throughout 6 target DNA concentrations. The lower detection limit based on plasmid DNA and the polymerase chain reaction products was 100 copies, whereas that obtained with genomic DNA reached up to 10 copies; this last system turned to be the most susceptible. CONCLUSIONS: real-time polymerase chain reaction system, standardized for the three standard DNA proved to be a rapid, specific and highly sensitive system for better diagnosis, and for the development of studies on the pathogenesis of human herpesvirus 8 infection in Cuba.

Keywords : human herpes virus 8; Karposi´s sarcoma; real-time polymerase chain reaction; Cuba.

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