Monterey Bay Aquarium Research Institute, 7700 Sandholdt Road, Moss Landing, California 95039 USA.
Mol Ecol Resour. 2008 May;8(3):540-50. doi: 10.1111/j.1471-8286.2007.02021.x.
Knowledge of the temporal and spatial abundance of invertebrate larvae is critical to understanding the dispersal capabilities and recruitment potential of marine and aquatic organisms. Traditional microscopic analyses are time-consuming and difficult given the diversity of larval species and a frequent lack of discriminating morphological characteristics. Here, we describe a sensitive rRNA targeted sandwich hybridization assay (SHA) that uses oligonucleotide probes to detect and enumerate the larvae of invasive green crabs (Carcinus maenas), native blue mussels (Mytilus), native barnacles (Balanus) and polychaetes (Osedax and Ophelia) that occur in the Monterey Bay National Marine Sanctuary, California. Laboratory-based assays demonstrate specificity, high sensitivity, and a quantitative response to cultured samples from three of the target organisms. Oligonucleotide probes were then printed in arrays on nitrocellulose membranes and deployed in our robotic Environmental Sample Processor (ESP) to detect larvae in situ and autonomously. We demonstrate that the SHA-detection method and ESP robot can be used for near real-time, in situ detection of larval species in the marine environment.
了解无脊椎动物幼虫的时空丰度对于理解海洋和水生生物的扩散能力和繁殖潜力至关重要。传统的显微镜分析方法耗时且困难，因为幼虫物种的多样性和经常缺乏有区别的形态特征。在这里，我们描述了一种灵敏的 rRNA 靶向夹心杂交分析 (SHA) ，该方法使用寡核苷酸探针来检测和计数入侵绿蟹 (Carcinus maenas) 、本地贻贝 (Mytilus) 、本地藤壶 (Balanus) 和多毛类动物 (Osedax 和 Ophelia) 的幼虫，这些幼虫存在于加利福尼亚州蒙特雷湾国家海洋保护区。基于实验室的分析表明，该方法具有特异性、高灵敏度和对三种目标生物培养样本的定量反应。然后，寡核苷酸探针被打印在硝酸纤维素膜上的阵列中，并在我们的机器人环境样品处理器 (ESP) 中自动部署，以原位检测幼虫。我们证明 SHA 检测方法和 ESP 机器人可用于海洋环境中实时、原位检测幼虫物种。
