A protocol for detection of mitochondrial DNA deletions: characterization of a novel deletion.

Abstract

OBJECTIVES:

To develop a protocol capable of identifying deletions in mitochondrial DNA and use it to identify the breakpoints of a mtDNA deletion in a patient with chronic progressive external ophthalmoplegia (CPEO).

DESIGN AND METHODS:

Deletions in mtDNA were identified by a combination of long range PCR and Southern blotting. The precise breakpoints were determined by automated DNA sequencing.

RESULTS:

A series of DNA samples from patients with suspected mitochondrial disease was subjected to a protocol, which combines long range PCR and Southern blotting. We found a unique deletion in a patient with CPEO and we identified the precise location of this deletion through DNA sequencing.

CONCLUSIONS:

Long range PCR has the advantages of speed, minimal samples requirements, and sensitivity. Southern blotting is better able to evaluate heteroplasmy and detect duplications. We suggest a protocol that enables us to identify precisely the breakpoints in a unique mutation of mtDNA in a patient with CPEO.

• Authors: M B Coulter-Mackie, D A Applegarth, J R Toone, L Gagnier
• Journal: Clinical biochemistry (Clin Biochem) Vol. 31 Issue 8 Pg. 627-32 (Nov 1998) ISSN: 0009-9120 [Print] United States
• PMID: 9876894 (Publication Type: Comparative Study, Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• DNA, Mitochondrial

Topics

• Adolescent
• Blepharoptosis (genetics)
• Blotting, Southern (methods)
• DNA, Mitochondrial (analysis)
• Humans
• Kearns-Sayre Syndrome (genetics)
• Male
• Mitochondrial Encephalomyopathies (genetics)
• Ophthalmoplegia (genetics)
• Ophthalmoplegia, Chronic Progressive External (genetics)
• Polymerase Chain Reaction (methods)
• Sensitivity and Specificity
• Sequence Deletion