超声波细胞粉碎机在高通量测序片段化 DNA 中的应用

摘要

Objective To investigate the optimal conditions of DNA fragmentation using a ultrasonic cell disrupter in high-throughput sequencing. Methods Under the conditions of ice bath,and of the diverse ul-trasonic power,gap/ultrasonic duration,ultrasonic time,and continuous operating time of the instrument,the DNA was broken with the ultrasonic cell disrupter and then analyzed using agarose gel electrophoresis and the Agilent 2100 expert_High Sensitivity DNA Assay to determine the effects of both DNA fragmentation and hy-droxyl methylation immune coprecipitation. Results Under the conditions of ice bath and ultrasonic power of 280W,gap/ultrasonic duration of 5s/5s,ultrasonic time of 18 min,and continuous operating time of the in-strument of 90min,we could harvest the genomic DNA fragment of 200 ~ 500 bp for the needs of high -throughput sequencing. Conclusion Using the ultrasonic cell disrupter under the optimized conditions,we can yield the repeatable DNA fragments which can be used to library construction for high-throughput sequen-cing.%目的：探讨超声波细胞粉碎机在高通量测序中应用的最佳 DNA 片段化条件。方法 DNA 样品在冰浴条件下采用不同的超声功率、单次间隙时间和超声时间、工作时间(单一样品超声破碎时间)、仪器连续工作时间,通过琼脂糖凝胶电泳和2100生物分析仪分别检测基因组 DNA 片段化和羟甲基化免疫共沉淀效果。结果超声功率280 W、间隙时间/超声时间5 s/5 s、总工作时间18 min,仪器连续工作时间不超过90 min,超声波处理时样品始终保持冰浴环境可获得高通量测序需要的、长度主要集中在200~500 bp 之间的 DNA 片段。结论在我们优化的实验条件下,利用超声波细胞粉碎机可以得到重复性较好的基因组 DNA 片段,可满足高通量测序文库构建中对 DNA 片段质量的要求。