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铜鱼(Coreius heterodon)作为长江中、上游的重要经济鱼类是研究三峡大坝阻隔效应的重要材料之一。然而,过去的铜鱼标本都保存在福尔马林溶液中,有必要探讨从福尔马林固定的铜鱼标本中有效提取基因组DNA的方法以及这些DNA用于微卫星和线粒体分析的可行性。本实验通过改进的酒精梯度浸泡法去除标本中的甲醛,然后用酚-氯仿抽提法成功地提取到了铜鱼标本的基因组DNA;设计引物后进行了线粒体和微卫星的PCR扩增,扩增产物经银染检测。微卫星扩增结果显示只有部分个体可以扩出目的带,而线粒体控制区部分区段在所有个体中均能稳定重复的扩出;mtDNASSCP分析显示带型一致。结果表明,福尔马林固定的铜鱼标本可以被用来开展短片段的扩增和遗传变异分析等方面的相关研究。
Coreius heterodon, one of the important economic fish in the middle and upper reaches of the Yangtze River, is one of the important materials for studying the barrier effect of the Three Gorges Dam. However, the past copperfish specimens were preserved in formalin solution, it is necessary to explore the method of extracting genomic DNA from formalin fixed copperfish specimens and the feasibility of using these DNA for microsatellite and mitochondrial analysis . In this experiment, formaldehyde was removed by the improved alcohol gradient immersion method, then the genomic DNA of the copper fish specimen was successfully extracted by phenol-chloroform extraction method. After the primer was designed, the mitochondrial and microsatellite PCR amplification and amplification The product was detected by silver staining. The result of microsatellite amplification showed that only some of the individuals could expand the target band, while some of the mitochondrial control region could stably and repeatedly expand in all the individuals. The mtDNASSCP analysis showed that the bands were consistent. The results show that formalin fixed copperfish specimens can be used to carry out short fragment amplification and genetic variation analysis and other related studies.