Tokuda Y, Nakamura T, Satonaka K, Maeda S, Doi K, Baba S, Sugiyama T
Department of Pathology, Kobe University School of Medicine, Japan.
J Clin Pathol. 1990 Sep;43(9):748-51. doi: 10.1136/jcp.43.9.748.
The mechanism of DNA degradation and its clinical applications were examined. When purified lambda phage and extracted liver DNA were fixed in phosphate buffered formaldehyde, the DNA did not degrade, but there was incomplete digestion with endonuclease. Rat liver tissues were fixed under various conditions and DNA extracted. Immediate fixation with buffered formaldehyde at low temperature, or the addition of EDTA to buffered formaldehyde blocked the DNA degradation. Analysis of pulsed field gel electrophoresis also showed that DNA was degraded before extraction. These results suggest that tissue nuclease has an important role in DNA degradation in tissue. Furthermore, formaldehyde fixation at low temperature, which may take time and which decreases slightly the staining capacity, is useful for the extraction of intact DNA. For clinical application, the detection of provirus was examined. Genomic DNA was extracted from a necropsy sample of adult T cell leukaemia fixed in formaldehyde; human T cell leukaemia virus type-I (HTLV-I) provirus was successfully detected by Southern blotting.
研究了DNA降解的机制及其临床应用。当纯化的λ噬菌体和提取的肝脏DNA固定在磷酸盐缓冲甲醛中时，DNA没有降解，但用核酸内切酶消化不完全。将大鼠肝脏组织在各种条件下固定并提取DNA。在低温下立即用缓冲甲醛固定，或在缓冲甲醛中添加EDTA可阻止DNA降解。脉冲场凝胶电泳分析也表明，DNA在提取前就已降解。这些结果表明，组织核酸酶在组织中的DNA降解中起重要作用。此外，低温甲醛固定虽然可能耗时且染色能力略有下降，但对完整DNA的提取很有用。对于临床应用，检测了前病毒。从用甲醛固定的成人T细胞白血病尸检样本中提取基因组DNA；通过Southern印迹法成功检测到I型人类T细胞白血病病毒（HTLV-I）前病毒。
