生物素标记探针检测外周血白细胞端粒长度的应用研究

A Biotin-Labeled Probe to Measure Telomere Length in Peripheral Leukocytes

目的建立生物素标记测定端粒DNA长度的方法,探讨其法医学应用价值.方法基因组DNA经RsaI和HinfI限制性内切酶消化,0.8%琼脂糖凝胶电泳,Southern印迹转移,以5'末端生物素标记寡核苷酸(TTAGGG)3为探针进行杂交,化学发光检测DNA谱带,与DNA标准分子量比较计算端粒长度.结果通过严格控制印迹转移条件、探针的工作浓度、杂交温度等主要影响因素,获得了比较好的杂交条带.结论生物素标记探针检测端粒DNA长度方法稳定、可靠,无放射污染,杂交信号明显.

Objective To measure telomere length by hybridization with biotin-labeled probe and study its application in forensic practice. Methods Genomic DNA was digested with restriction enzyme Rsa Ⅰ and Hinf Ⅰ, the products were separated by 0.8% agarose gel electrophoresis followed by Southern blotting with 5＇ terminal labeled oligonucleotide （TTAGGG） 3 probe and chemiluminescence detection. The mean telomere length was calculated by comparing with standard kilobase sized marker. Results The hybridization bands were showed by stringently controlling the transfer condition, the probe working concentration and hybridization temperature （55 ℃ ）. Conclusion Hybridization with biotin-labeled probe gives strong hybridization signal and is non-radioactive, and is reliable and stable to determinate telomere length.