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Successful quantification of cytomegalovirus DNA by competitive PCR and detection with capillary electrophoresis.

Abstract：

:Human cytomegalovirus (HCMV) is responsible for severe infections in immunocompromised patients. Viral load has recently been identified as one of the major risk factors for subsequent development of HCMV disease. In this context, we developed a protocol allowing rapid, sensitive and precise quantification of HCMV DNA using competitive PCR run to saturation. Long primers were used for amplification, and internal DNA standard was constructed by PCR, with a primer inducing formation of a loop on the target sequence. The obtained fragment differed from the wild one (142 bp) by 6 bp. Quantitative analysis of PCR-amplified HCMV DNA was carried out using an original system combining capillary gel electrophoresis and u.v. detection. This procedure was evaluated on renal transplant recipients, and the results of quantitative PCR were compared with those of viraemia, qualitative DNAemia and HCMV-related symptoms. High levels of HCMV DNA were associated with HCMV-related symptoms, and in all cases a significant decrease of viral load was observed following DHPG treatment. Competitive PCR with capillary electrophoresis detection appears to provide a sensitive quantification method for HCMV DNA in leukocytes and is easily adaptable to routine laboratory use.

journal_name

Mol Cell Probes

journal_title

Molecular and cellular probes

authors

Poirier-Toulemonde AS,Imbert-Marcille BM,Ferré-Aubineau V,Besse B,Le Roux MG,Cantarovich D,Billaudel S

doi

10.1006/mcpr.1996.0071

subject

Has Abstract

pub_date

1997-02-01 00:00:00

pages

11-23

issue

1

eissn

0890-8508

issn

1096-1194

pii

S0890-8508(96)90071-7

journal_volume

11

pub_type

杂志文章

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