Quantitation of mitochondrial DNA in human lymphoblasts by a competitive polymerase chain reaction method (Meeting abstract).

Important note: Information in this article was accurate in 1995. The state of the art may have changed since the publication date.

Quantitation of mitochondrial DNA in human lymphoblasts by a competitive polymerase chain reaction method (Meeting abstract).

Proc Annu Meet Am Assoc Cancer Res; 35:A3458 1994. Unique Identifier : AIDSLINE ICDB/95605143
Hao Z; Cooney DA; Zhan Q; Sreenath A; Stowe E; Johns DG; LMC, NIH, Bethesda, MD 20892

Abstract: A quantitative competitive polymerase chain reaction method has been developed to measure the copy numbers of mitochondrial DNA (mtDNA) in MOLT-4 cells. A competitive DNA template was synthesized by PCR, the product containing a central fragment of HIV DNA flanked by 20-mer primer sequences identical to portions of the mtDNA D-Loop. Copy numbers of this product were determined. For the assay, increasing known copy numbers of the competitive template were added to samples containing total cellular DNA. Two QC-PCR products were generated: (1) a 181 bp fragment derived from the D-Loop region of mtDNA (bp 6-187); and (2) a 155 bp fragment derived from the competitive DNA template. Absolute quantitation was achieved by radiometric comparison of the relative amounts of the two products. Varying amounts of the competitive template were analyzed with a fixed quantity of total cellular DNA; copy numbers of mtDNA were 73 +/- 11 copies/cell. At a low concentration of a known inhibitor of mtDNA synthesis, 2',3'-dideoxycytidine (50 nM for 9 days), mtDNA was found to decrease from 73 to 2 copies/cell. The method is of general applicability to the study of mtDNA synthesis inhibitors.

Keywords: Cells, Cultured DNA Primers DNA, Mitochondrial/*METABOLISM Human Lymphocytes/*METABOLISM Polymerase Chain Reaction Templates ABSTRACT
950330
M9530830

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