2种保存方法对五唇兰基因组DNA提取质量的影响

Effects of Two Sample Preserving Methods on Genomic DNA Extraction Quality of Phalaenopsis pulcherrima(Orchidaceae)

采用改良CTAB法研究经硅胶干燥法和保鲜法保存的五唇兰叶片DNA降解规律,并比较两种保存方法叶片DNA的保存时间,为叶片的野外远距离采集提供指导。结果显示,采取硅胶干燥法进行保存的五唇兰叶片DNA和SOD以及POD的含量下降迅速,第1天DNA的产率已经下降到65.4 ng/g,ISSR引物可以检测出7条清晰的条带,第2天DNA则降解完全,SOD和POD含量2 d内快速较少,第2天分别降到68%和27%;采取叶片保鲜法保存的五唇兰叶片DNA一周内保存完好,产率较高,为71 ng/g左右,ISSR引物可以检测出7条清晰的条带,7 d之后DNA开始降解,ISSR引物扩增出的条带变得模糊,随着叶片衰老进程的加剧以及叶片开始出现腐烂等性状,DNA降解逐渐加速,第13天降解完全,SOD和POD含量的变化呈现出与DNA产率一致的趋势,先上升后下降,在第9天达到峰值。叶片DNA的降解规律和衰老规律呈现出一致性,离体五唇兰叶片DNA降解主要原因来自于叶片衰老的加剧以及期间产生的有毒物质的积累,野外远距离采集五唇兰叶片可采用保鲜法保存以防止DNA的快速降解。

A modified CTAB method was used to research the DNA degradation rule of P. pulcherrima leaves saved with silica gel and the low temperature fresh-keeping method respectively. The results showed that DNA degradation of leaves saved with silica gel was rapid, the DNA production rate after one day dropped to 65.4 ng/g, seven clear bands were detected using ISSR primers, and DNA degraded completely after two days; However, DNA of P. pulcherrima leaves saved with low temperature fresh-keeping method could be preserved for one week, and the yield was higher, about 71 ng/g, ISSR primers could detect seven clear bands, after 7 days DNA began to degrade and PCR bands became blurry The rule of DNA degradation and senescence showed consistency. The main reasons for degradation of leaf DNA were intensification of senescence and the accumulation of toxic substances produced during the period. Leaves of P. pulcherrima sampled in the wild can take the method of low temperature to keep fresh.