Abstract: Spore cysts of parasite Thelohanellus wuhanensis collected from diseased juvenile allogynogenetic crucian carp Carassius auratus gibelio were preserved in 100%ethanol，70%ethanol，2.5%glutaraldehyde，10%formalin，Bouin’s solution，Carnoy’s solution or Müller’s solution for 50 days to explore effects of different preservative methods on DNA extraction，conventional PCR amplification and nested PCR amplification of the parasite，and fresh spores were used as control group.The spores from the experimental groups and control group were counted by blood count plates and adjusted by distilled water with same concentration in each group.DNA extractions，conventional PCR amplifications and nested PCR amplifications of spores with equal number from different groups were made and their products were evaluated through agarose gel electrophoresis to explore the effects of different preservations.The results showed that 100%ethanol and 70%ethanol preservations do not impact DNA extractions，conventional PCR amplifications and nested PCR amplifications，with the same bright and consistent specific electrophoretic bands in agarose gel electrophoresis as those in the fresh spores in the control group，indicating that 100% ethanol and 70%ethanol can be used as the preservative solutions for spores.However，2.5%glutaraldehyde and Bouin’s solution preservations showed influences on the DNA extractions，with specific electrophoresis bands only in the products of the conventional and nested PCR amplification，and thus 2.5%glutaraldehyde and Bouin’s solution can be used as the preservative solutions of spores for the conventional and nested PCR amplifications.It was found that 10%formalin and Müller’s solution preservations led to influences on the DNA extractions and the conventional PCR amplifications，with specific electrophoresis bands only in the nested PCR amplifications.It is conferred that 10%formalin and Müller’s solution can only be used as the preservative solutions of spores for nested PCR amplification.There was no electrophoresis bands from the products of DNA extraction as well as the conventional and nested PCR amplifications in Carnoy’s solution preservation，which cannot be used as the preservative solutions of spores for molecular biology analyses associated with nucleic acid.The findings indicate that except for 100%and 70%ethanol，other preservative methods have different degree of effects on DNA extraction，conventional PCR amplification and nested PCR amplification of the spores.
陆宏达, 刘俊杰, 岳蒙蒙, 操艮萍, 李振伟. 不同保存方法对武汉单极虫DNA提取和PCR扩增的影响[J]. 大连海洋大学学报, 2016, 31(3): 246-251.
LU Hong-da, LIU Jun-jie, YUE Meng-meng, CAO Gen-ping, LI Zhen-wei. Effects of different preservation methods on DNA extraction and PCR amplification in parasite Thelohanellus wuhanensis. Journal of Dalian Ocean University, 2016, 31(3): 246-251.
2016, Vol. 31 
