摘要
在纤毛虫的分子生物学研究中,单细胞基因组DNA的微量提取是后续基因分析的关键。本研究以海洋纤毛虫红色伪角毛虫为材料,采用直接取完整虫体、裂解法以及3种改良后的试剂盒微量提取DNA方法,分别提取了新鲜样品与4种不同方法保存样品的基因组DNA,比较了9种组合在PCR产物质量、提取所需时间、成本及便捷性方面的优劣,并进一步比较、讨论和探索了4种样品保存方法和5种DNA微量提取方法在纤毛虫单细胞基因组PCR扩增中的可行性。本工作旨在为后续的纤毛虫分子生物学研究提供可资借鉴的技术路线和方法。
DNA micro-extraction is one of the bottlenecks for molecular investigations on ciliated protozoa since most ciliates are difficult to be cultured. Using Pseudokeronopsis rubra, a marine hypotrichous ciliate, as sample, five DNA micro-extraction methods （viz. three improved DNA micro-extraction kit methods, lysis method and using single cell directly） for living single cell and four sample preservation methods （viz. 95% alcohol with 100μM EDTA, two kits） are performed. By comparing these nine combi- nation methods for DNA micro-extraction and sample preservation, following conclusions could be drawn： REDExtract-N-AmpTM Tissue PCR Kit and lysis method are the best ones for living samples, and DNeasy Blood ~ Tissue Kit and Indicating Whatman FTA Elute Cards are suggested for DNA micro-ex- traction of environmental samples, which could not be treated in time.
出处
《中国海洋大学学报（自然科学版）》
CAS
CSCD
北大核心
2012年第7期60-63,共4页
Periodical of Ocean University of China
基金
国家自然科学基金项目(41006098
31071893)
广东高校优秀青年创新人才培养计划(LYM10060)资助
关键词
纤毛虫
红色伪角毛虫
单细胞DNA微量提取
样品保存
基因扩增
ciliates
Pseudokeronopsis rubra
DNA micro-extraction of single cell
sample preservation
gene amplification
