[Detection of mtDNA 11778 (G-->A) point mutation in a family with Leber's hereditary optic neuropathy by site-specific polymerase chain reaction].
OBJECTIVE
To find a simple, fast, accurate, and quantitative PCR-based method for mutation detection, so as to identify mitochondrial DNA 11778 G-->A point mutation in patients with Leber's hereditary optic neuropathy (LHON).


METHODS
On the basis of sequencing of mtDNA from LHON proband, M primer for mutation and N primer for normal were designed to be coupled with reverse primer respectively. Specific PCRs were done on an amplifying condition with high stringency such as a well controlled annealing temperature, low Mg2+ concentration and less thermal cycles. The objective pedigree includes 10 individuals, were against 40 normal control persons.


RESULTS
Different ratios of indicative mtDNA 11778A-->G mutation were checked out from the proband, affected maternal members and a 10 year-old boy (up to now no appearance yet), whereas not appeared on normal spouses, paternal offsprings in the family, neither did on 40 controls.


CONCLUSIONS
This site-specific PCR method is a kind of general mutation analysis way, without the restriction of existence of endonuclease site. It can be applied for the gene diagnosis of known-mutation hereditary diseases such as LHON.
Niu
Shu-lan
SL
Central Laboratory, First Hospital of Peking University, Beijing 100034, China.
Zhang
Ying
Y
Xu
Yu-feng
YF
Bu
Ding-fang
DF
Ren
Ze-qin
ZQ
Wang
Shao-yu
SY
Liu
Guo-hua
GH
Qi
Yu
Y
D000328:Adult; D004272:DNA, Mitochondrial / Q000235:genetics*; D005260:Female; D006801:Humans; D008297:Male; D029242:Optic Atrophy, Hereditary, Leber / Q000235:genetics*; D010375:Pedigree; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000379:methods
D004272:DNA, Mitochondrial