贝类单孢子虫二温式PCR检测方法的建立及初步应用

• 作 者: 谢丽基 谢芝勋 刘加波 庞耀珊 邓显文 谢志勤 范晴
• 机 构: 广西壮族自治区兽医研究所 广西畜禽疫苗新技术重点实验室,
• 出 处: 黑龙江畜牧兽医
• 发表日期: 2013年13期
• 关键词: 单孢子虫 PCR检测方法 河弧菌 特异性扩增带 溶藻弧菌 副溶血弧菌 扩增条件 菲律宾蛤仔 特异性试验 试验设计

为了建立检测贝类单孢子虫的二温式PCR方法,试验根据GenBank中单孢子虫的基因保守序列设计了1对特异性引物,并对二温式PCR扩增条件进行优化。结果表明:用二温式PCR方法对单孢子虫模板进行扩增,得到与试验设计相符的333 bp特异性扩增带,而对折光马尔太虫、派琴虫、副溶血弧菌、溶藻弧菌和河弧菌等病原体进行扩增,结果全为阴性;该方法最低能检测到1 fg的单孢子虫质粒DNA。

In order to establish a two-temperature PCR method for detecting C. monocytogenes, a pair of specific primers was designed based on the gene sequence of the single sporozoite gene in GenBank. The two-temperature PCR amplification conditions were optimized. The results showed that the amplification of the template of monosporidium was carried out by a two temperature PCR method, and the 333 bp specific amplification band corresponding to the experimental design was obtained. However, Vibrio and Vibrio cholerae and other pathogens were amplified, the results were all negative; the method can detect the lowest 1 fg of the sporozoite plasmid DNA.