可用于相关亚历山大藻(Alexandrium affine)检测的一个寡核苷酸探针

A OLIGONUCLEOTIDE PROBE FOR DETECTION OF ALEXANDRIUM AFFINE

针对分离自南海的相关亚历山大藻,在进行核糖体大亚基DNA(LSU rDNA)全序列测序及比对的基础上,设计了针对其核糖体大亚基RNA(LSU rRNA)D10区的一个新的荧光素(FITC)标记寡核苷酸探针,并建立了相应的荧光原位杂交检测方法。在GenBank数据库中,BLAST检索的结果显示所设计的探针具有较高的特异性。杂交实验的结果也表明,这条探针对相关亚历山大藻的标记具有特异性,且标记效率可以达到100%,标记信号清晰明亮。本研究结果为相关亚历山大藻的监测及其种群动态研究提供了方法学依据,也为其它微藻特异性探针的设计提供了新的思路。

Many species in Genus Alexandrium were important harmful-algal-bloom causative species for their capability in producing paralytic shellfish poisons （PSPs）. Alexandrium affine, however, does not produce PSPs, but it can still do harm to the growth, reproduction and development of organisms such as rotifer, cladoceran, crustacean and larvae of molluscan, through a kind of unidentified toxins. Using a strain of A. affine （AC-1） isolated from the South China Sea of China, we designed a new oligonucleotide probe （AF-1） based on the analysis of its sequence information of the large sub-unit ribosomal RNA （LSU rRNA） gene and the comparison of the LSU rRNA gene sequence with related species. The probe, specifically targeted on the D10 region of the LSU rRNA, was labeled with fluorescein isothiocyanate （FITC）. A fluorescence in situ hybridization （FISH） method for the “whole-cell” identification of A. affine was also established. A BLAST analysis of the designed probe in the GenBank database found that the probe AF-1 had a high specificity in identi-fication of A. affine. The FISH experiment also confirmed that the probe AF-1 could specifically label the cells of A. affine, not the other algal cells used in the experiment. The labeling efficiency of target cells could reach 100%, with bright green signal under fluorescence microscope. These results laid a technical basis for monitoring and studying of A. affine and its population dynamics, and offered a new idea for the designation the specific probes for other algae.