基于流式细胞分析技术的茯苓基因组大小测定

Flow Cytometric Analysis of Nuclear DNA Content in Poria cocos

目的:茯苓是担子菌多孔菌科的著名传统中药。本研究的目的在于测定茯苓的基因组大小并考察了其倍性与基因组的关系。方法:真菌材料的前处理参考文献报道[1],并用DNA特异性染料碘化丙啶进行染色,通过调节离心转速、染色时间等条件对样品进行核分离的优化。DNA含量用流式细胞仪进行分析,通过比较茯苓与内参黑曲霉G0/G1期峰比值计算得出。结果:茯苓的基因组大小测定为55.79±3.03Mb,或相对DNA含量为0.12±0.01pg(1pgDNA=0.965×109bp)。在结果中可以检测出黑曲霉和茯苓的G2/M期峰。结论:本研究探索了用流式细胞仪分析茯苓基因组大小的方法,所得数据可为基因组学研究提供了参考。

Poria cocos, which belongs to Polyporaceae in basidiomycetes, is a well-known Traditional Chinese Medicine （TCM）. The present study aims to estimate the DNA content as well as the relationship between ploidy and DNA content. The fungal material was prepared according to a protocol adopted from Jaroslav Dolezel et al. [1] and stained with propidium iodide mized by adjusting centrifugal （PI） which is a nucleic acid dye. For this study, a nuclei isolating method was optiforce and staining period. The DNA quantity was quantified by flow cytometry and calculated as the ratio of mean flurescence of the G0/G1 peak between Poria cocos and AspergiUus niger. The Aspergillus niger was used as an internal reference. The estimated genome size of Poria cocos is 55.79±3.03 Mb, or 2C DNA about 0.12±0.01 pg （lpg DNA =0.965×10^9 bp）. The peak of GJM of Poria cocos and Aspergillus niger could be detected. In summary, the present study described a modified method of genome size analysis by using flow cytometry in Poria cocos. The data available of DNA content obtained in this study is helpful to the future genome sequencing research.