Rapid detection of mitochondrial DNA polymorphism by photoactivatable biotin.
A new method for the detection of mitochondrial (mt)DNA polymorphism by agarose gel electrophoresis is presented. In this method, after mtDNA digestions with endonucleases, the mtDNA fragments are labeled directly with photoactivatable biotin in the digestion buffer, separated by agarose gel electrophoresis, transferred to a nylon membrane and detected enzymatically using streptavidin-alkaline phosphatase, Nitroblue tetrazolium (NBT) and 5-bromo-4-chloro-3-indolylphosphate (BCIP). This method is useful for the detection of mtDNA digestion patterns by agarose gel electrophoresis, especially when enough purified mtDNA cannot be obtained and the radioisotope cannot be handled because of several restrictions.
Tsuchida
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Department of Legal Medicine and Human Genetics, Jichi Medical School, Tochigi-Ken, Japan.
Ikemoto
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D000469:Alkaline Phosphatase; D000818:Animals; D001426:Bacterial Proteins; D001710:Biotin; D004272:DNA, Mitochondrial / Q000032:analysis* / Q000302:isolation & purification; D015245:Deoxyribonuclease BamHI; D015246:Deoxyribonuclease EcoRI; D004285:Dogs; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D007211:Indoles; D007668:Kidney / Q000737:chemistry; D009580:Nitroblue Tetrazolium; D012150:Polymorphism, Restriction Fragment Length; D019809:Streptavidin
D001426:Bacterial Proteins; D004272:DNA, Mitochondrial; D007211:Indoles; D009580:Nitroblue Tetrazolium; C035455:5-bromo-4-chloro-3-indoxyl phosphate; D001710:Biotin; D019809:Streptavidin; D015245:Deoxyribonuclease BamHI; D015246:Deoxyribonuclease EcoRI; D000469:Alkaline Phosphatase