松墨天牛成虫标本保存及其DNA提取质量比较

Comparison of the quality of genomic DNA extracted from adult specimens of Monochamus alternatus preserved by different methods

【目的】探讨适合DNA提取的天牛成虫标本保存方法。【方法】采用SDS-蛋白酶K消化法对液氮中冷冻保存、无水乙醇-20℃冷冻保存、无水乙醇室温保存和干标本室温保存且保存时间在2年以上的松墨天牛Monochamus alternates Hope成虫标本基因组DNA进行提取,并对不同保存方式提取的DNA样本进行了质量比较和分析。【结果】在上述常见的松墨天牛成虫标本4种保存方式中,以液氮中冷冻保存效果最佳,其次为无水乙醇-20℃冷冻保存,插针干标本室温保藏效果最差。利用昆虫线粒体基因CO I和CO II的通用引物从上述DNA中均能够成功扩增出目的片段,测序结果证实扩增片段符合预期。【结论】液氮和无水乙醇-20℃冷冻保存适合松墨天牛成虫标本长期保存,且不影响后续的PCR扩增和测序。

[Objectives] To find which method of preserving Monochamus alternatus specimens produced the best quality genomic DNA. [Methods] Genomic DNA from Monochamus alternatus adults that had been preserved using different methods （liquid nitrogen, 100% ethanol at -20℃, 100% ethanol at room temperature and museum specimens） for more than 2 years, was extracted using the SDS-proteinase K method and the quality of the extracted genomic DNA compared. [Results] The best quality genomic DNA was obtained from specimens that had been stored in liquid nitrogen, and the next best from those that had been preserved in 100% ethanol at -20℃. [Conclusion] The best quality DNA was extracted from specimens that had been frozen in liquid nitrogen or preserved in 100% ethanol at -20℃. The DNA samples obtained from such specimens are suitable for PCR amplification and sequencing.