Comparison of three methods for isolation of nucleic acids from membranate inner ear tissue of rats

• Published in: Chinese medical journal Vol. 119; no. 12; pp. 986 - 990
• Main Authors: KONG, Wei-jia, WANG, Ying, WANG, Qiong, HAN, Yue-chen, HU, Yu-juan
• Format: Journal Article
• Language: English
• Published: China Department of Otorhinolaryngology,Union Hospital of Tongji Medicial College,Huazhong University of Science and Technology,Wuhan 430022,China 20.06.2006
• Subjects: Animals; DNA - isolation & purification; DNA, Mitochondrial - isolation & purification; Ear, Inner - chemistry; Polymerase Chain Reaction; Rats; Rats, Wistar; RNA - isolation & purification; 聚合酶链反应; 薄膜组织; 隔离方法; nuclear DNA; RNA; mitochondrial DNA; polymerase chain reaction
• ISSN: 0366-6999; 2542-5641
• DOI: 10.1097/00029330-200606020-00004

Background Mitochondrial DNA mutations have been found in sensorineural deafness. The aim of this study was to compare three methods for extraction of nucleic acid from membranate inner ear tissue of rats. Methods Alkaline denaturation, a conventional phenol-chloroform method and Trizol reagent were respectively used to extract the slight nucleic acid from membranate inner ear tissue of rats. We assessed the amount and quality of nucleic acid using a UV-spectrometer and polymerase chain reaction （PCR）. Results The yield and purity （OD260/OD280） of DNA from inner ear tissue using the phenol-chloroform method was the highest of the three methods. Mitochondrial DNA （mtDNA） fragment can be amplified by PCR from nucleic acid prepared by all methods, while no nuclear DNA （nDNA） fragment can be amplified by method of alkaline denaturation. Both nuclear and mitochondrial genes could be amplified by reverse transcriptional PCR from the RNA prepared by Trizol reagent. Conclusion Adequate amount and high-quality of mtDNA, nDNA and RNA were obtained from unilateral membranate inner ear tissue of rats. Method of alkaline denaturation could be chosen when mtDNA without nDNA was needed, while phenol-chloroform method was suitable for extracting total DNA （including nDNA and mtDNA）： method with Trizol reauent was suitable for extractinu total RNA and total DNA.