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[Extraction, amplification, recombination and sequencing of the mitochondrial DNA from celloidin embedded human
Objective:
To establish an analytical method of DNA extracted from human celloidin-embedded temporal bone sections by molecular biologic technique.
Methods:
Polymerase chain reaction (PCR) with different primers, special amplification methods or coorperated with enzyme restricted reactions was adapted to detect several types of genetic mutations from 9 human celloidin-embedded temporal bone sections. The pGEM-T vector was used for the recombination and cloning of the amplified DNA fragment.
Results:
The 135 bp fragments of normal mtDNA were detected from all of the 9 cases. The 316bp fragment related to mtDNA 4977bp deletion was also detected in two cases who suffered from presbycusis before death by the nested PCR. The result of sequencing confirmed the accuracy of PCR.
Conclusion:
The application of molecular biologic techniques in temporal bone research is important in improving the quality and value of retrospective study of ear diseases and has resulted in findings of the relevant mutant or pathogenetic genes in the ear diseases such as presbycusis, ototoxic deafness, otosclerosis and viral infections of the ear.