骨髓细胞固定液DNA提取与质量评价的实验研究

Study on the Genomic DNA Extraction from the Fixative Solution of Bone Marrow Cells and Its Quality Evaluation

目的利用骨髓染色体核型分析后剩余骨髓细胞固定液标本提取基因组DNA并评价DNA质量。方法将60例骨髓细胞固定液标本按保存时间的不同分为3组，同时另取新鲜骨髓标本（冻存于-80℃）20例，利用天根公司DNA提取试剂盒（离心柱型）提取基因组DNA，通过紫外分光光度仪检测DNA浓度与纯度、DNA凝胶电泳观察DNA条带、聚合酶链反应检测有无不同片段大小的目的基因的扩增以评价DNA质量。结果骨髓细胞固定液标本及新鲜骨髓（冻存）标本所提取的基因组DNA的A260／A280比值分别为1．93±0．03、1．92±0．04；两者纯度比较，差异无统计学意义（P〉0．05）；不同保存时间（〈3年、3～5年、〉5年）的骨髓细胞固定液标本所提取DNA的A260／A280比值分别为1．94±0．03、1．91±0．03、1．93±0．04，其纯度差异无统计学意义（P〉0．05）。上述实验标本均能通过PCR反应扩增出不同片段大小的目的条带。结论利用染色体核型分析后剩余的骨髓细胞固定液标本可以提取出质量合格的基因组DNA，其可作为分子生物学相关研究的标本来源。

Objective To extract genomic DNA from the fixative solution of bone marrow cells after the analysis of chromosome karyotyping and detection of the quality. Methods We extracted genomic DNA from the fixative solution of bone marrow cells and the samples of fresh bone marrow （frozen in - 80℃ ） with TIANGEN kit. The concentration and purity were detected by ultraviolet spectrophotometer. The integrity of DNA was measured by agarose gel electrophoresis and polymerase chain reaction. Results The ratio of A260/ A280 of genomic DNA extracted from the fixative solution of bone marrow cells and the samples of fresh bone marrow were 1.93 ±0.03, 1.92 ± 0.04 respectively and the statistical significant difference of the DNA purity wasn＇t found between them. The ratio of A260/A280 of genomic DNA extracted from the three groups of fixative solution of bone marrow cells divided by different preservation time （ less than 3 years,3 to 5 years, more than 5 years） were 1.94 ± 0.03,1.91 ± 0.03,1.93 ±0.04 respectively and also the statistical difference didn＇t exist among them. All of the experimental samples could amplify the target fragments with different sizes. Conclusion The quality of genomic DNA from the fixative solution of bone marrow cells was qualified. The fixative solution of bone marrow cells can be used as the source of specimen for molecular biology research.