珍稀濒危植物翅果油树基因组大小的测定

Determination of Genome Size of a Rare and Endangered Plant Elaeagnus mollis Diels

本文的研究目的是通过流式细胞法来测定珍稀濒危植物翅果油树基因组的大小。研究采用基因组大小已知的中国沙棘作为参考植物。首先,通过两种温度浸种优化了两种植物种子萌发的条件,培养出幼苗。其次,比较了OttoI裂解液对翅果油树和中国沙棘幼苗叶片的细胞裂解效果,并通过徒手切片法观察了二者叶片的细胞大小。最终确定选用OttoI和OttoII细胞裂解液提取两种植物叶片的细胞核悬浮液并根据细胞大小确定了相同的细胞悬浮液上样量,同时用碘化丙啶进行细胞核染色,将处理好的样品上流式细胞仪进行细胞核DNA含量的测定。通过比较内参样品中国沙棘与待测样品翅果油树二者细胞核的G0/G1期峰值,计算出翅果油树的基因组大小。结果以中国沙棘为内参测定的翅果油树的基因组大小为1652.82 ±78.24 Mbp,或相对DNA含量为1.69 ±0.08 pg。研究结果为翅果油树基因组的测序等研究提供了数据参考。

The study aims to determine the genome size of a rare and endangered plant Elaeagnus mollis by using flow cytometry method. Hippophae rhamnoides ssp. sinensis that the genome size was known was selected as an internal reference plant. First, the conditions of seed germination of plants were optimized by soaking seeds in different temperatures water and the seeds were cultured in soil. Then the study compared the cracking effects of OttoI lysate on plant leaves of Elaeagnus mollis and Hippophae rhamnoides ssp. sinensis. Meanwhile, we observed the cell size by freehand sectioning. Finally, we extracted nuclei with OttoI and OttoII lysates and nuclei were stained with propidium iodide. Based on the observation both of cell size, we made sure the same leaves weight and sample loading. Then, these samples were detected by flow cytometry and their DNA contents were calculated as the ratio of mean fluorescence of the G0/G1 peak between Elaeagnus mollis and Hippophae rhamnoides ssp. sinensis. The results show that the determined genome size of Elaeagnus mollis is 1662.6 ±78.24 Mbp, or relative DNA content is 1.70 ±0.08 pg. The results provide a data reference for future research of genome sequencing of Elaeagnus mollis.