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Clinical Chemistry 44: 2088-2093, 1998;
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(Clinical Chemistry. 1998;44:2088-2093.)
© 1998 American Association for Clinical Chemistry, Inc.

Molecular Diagnostics and Genetics

New sensitive method for the detection of the A3243G mutation of human mitochondrial deoxyribonucleic acid in diabetes mellitus patients by ligation-mediated polymerase chain reaction

Michiyo Urata1, Machiko Wakiyama1, Masanori Iwase2, Makoto Yoneda3, Sachiko Kinoshita1, Naotaka Hamasaki1, and Dongchon Kang1,a

1 Clinical Laboratory, Kyushu University Hospital, Fukuoka 812-8582, Japan.

2 Second Department of Internal Medicine, Kyushu University Faculty of Medicine, Fukuoka 812-8582, Japan.

3 Second Department of Internal Medicine, Fukui Medical University, Fukui 910-1193, Japan.
a Address correspondence to this author at: Department of Clinical Chemistry and Laboratory Medicine, Kyushu University Faculty of Medicine, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan. Fax 81-92-642-5772; e-mail kang{at}biochem2.med.kyushu-u.ac.jp.

An adenine-to-guanine mutation at nucleotide position (np) 3243 in the mitochondrial tRNALeu(UUR) gene is closely associated with various clinical phenotypes of diabetes mellitus. Because the mutation creates a new restriction site for the restriction enzyme ApaI, the mutation is usually detected and quantified by ApaI cleavage of the PCR products including np 3243. The sensitivity of the conventional method is, however, 5–10% heteroplasmy. The percentage of heteroplasmy of the mutation is usually highest in the affected tissues and is much lower in peripheral blood cells, which are used most frequently for the analysis. The sensitivity of the conventional method, however, is not sufficient to detect the mutation from peripheral blood cells. Utilizing ligation-mediated polymerase chain reaction, we have developed a feasible and sensitive method to detect 0.01% heteroplasmy of the 3243 mutation in peripheral leukocytes.




The following articles in journals at HighWire Press have cited this article:


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K. S. Lim, R. K. Naviaux, and R. H. Haas
Quantitative Mitochondrial DNA Mutation Analysis by Denaturing HPLC
Clin. Chem., June 1, 2007; 53(6): 1046 - 1052.
[Abstract] [Full Text] [PDF]

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R. Singh, S. Ellard, A. Hattersley, and L. W. Harries
Rapid and Sensitive Real-Time Polymerase Chain Reaction Method for Detection and Quantification of 3243A>G Mitochondrial Point Mutation
J. Mol. Diagn., May 1, 2006; 8(2): 225 - 230.
[Abstract] [Full Text] [PDF]

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 Clin. Chem.Home page
M. Urata, Y. Wada, S. H. Kim, W. Chumpia, Y. Kayamori, N. Hamasaki, and D. Kang
High-Sensitivity Detection of the A3243G Mutation of Mitochondrial DNA by a Combination of Allele-Specific PCR and Peptide Nucleic Acid-Directed PCR Clamping
Clin. Chem., November 1, 2004; 50(11): 2045 - 2051.
[Abstract] [Full Text] [PDF]

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K. Ohkubo, A. Yamano, M. Nagashima, Y. Mori, K. Anzai, Y. Akehi, R. Nomiyama, T. Asano, A. Urae, and J. Ono
Mitochondrial Gene Mutations in the tRNALeu(UUR) Region and Diabetes: Prevalence and Clinical Phenotypes in Japan
Clin. Chem., September 1, 2001; 47(9): 1641 - 1648.
[Abstract] [Full Text] [PDF]


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