Haley S T, Cavender J F, Murray T E
Department of Biology, Elizabethtown College, PA 17022, USA.
Biotechniques. 1999 Jan;26(1):88-91. doi: 10.2144/99261st01.
Alexandrium tamarensis is a toxigenic dinoflagellate found in coastal waters worldwide. A critical factor in alleviating the health and economic threats posed by this species is the development of a rapid and reliable method for detection. This study stream-lined a labor- and resource-intensive protocol for the isolation of A. tamarensis ribosomal DNA (rDNA). Subcultures of A. tamarensis were established in water samples from several coastal Atlantic sites. A commercial DNA isolation kit protocol for cultured cells was used for isolation of the dinoflagellate DNA. Samples were amplified by PCR using primers specific for a 700-bp sequence of A. tamarensis rDNA. It was determined that this method facilitated the detection of 10(-4) ng/microL of A. tamarensis DNA. Furthermore, the kit enabled A. tamarensis to be isolated from the water sources with little signal degradation. This is a valuable technique for the rapid detection of A. tamarensis, even before cell numbers are large enough for morphological identification.
塔玛亚历山大藻是一种产毒的甲藻，在世界各地的沿海水域均有发现。减轻该物种对健康和经济造成威胁的一个关键因素是开发一种快速可靠的检测方法。本研究简化了一种用于分离塔玛亚历山大藻核糖体DNA（rDNA）的耗费人力和资源的方案。从大西洋沿岸多个地点的水样中建立了塔玛亚历山大藻的亚培养物。使用针对培养细胞的商业DNA提取试剂盒方案来提取甲藻DNA。使用针对塔玛亚历山大藻rDNA 700碱基对序列的特异性引物通过PCR对样品进行扩增。结果表明，该方法有助于检测到10⁻⁴ ng/μL的塔玛亚历山大藻DNA。此外，该试剂盒能够从水源中分离出塔玛亚历山大藻，且信号几乎没有降解。即使在细胞数量不足以进行形态鉴定之前，这也是一种用于快速检测塔玛亚历山大藻的有价值技术。
