Chromosomal markers of the diversity and evolution of dinoflagellates are scarce because the genomes of these organisms are unique among eukaryotes in terms of their base composition and chromosomal structure. Similarly, a lack of appropriate tools has hindered studies of the chromosomal localization of 5S ribosomal DNA (rDNA) in the nucleosome-less chromosomes of dinoflagellates. In this study, we isolated and cloned 5S rDNA sequences from various toxin-producing species of the genus Alexandrium and developed a fluorescence in situ hybridization (FISH) probe that allows their chromosomal localization. Our results can be summarized as follows: 1) The 5S rDNA unit is composed of a highly conserved 122-bp coding region and an intergenic spacer (IGS), the length and sequence of which are variable even within strains. 2) Three different IGS types, one containing the U6 small nuclear RNA (snRNA) gene, were found among four of the studied species (A. minutum, A. tamarense, A. catenella and A. pacificum). 3) In all strains investigated by FISH (A. minutum, A. tamarense, A. pacificum, A. catenella, A. andersonii and A. ostenfeldii), 5S rDNA gene arrays were separate from the nucleolar organizer region, which contains the genes for the large 45S pre-ribosomal RNA. 4) One to three 5S rDNA sites per haploid genome were detected, depending on the strains/species. Intraspecific variability in the number of 5S rDNA sites was determined among strains of A. minutum and A. pacificum. 5) 5S rDNA is a useful chromosomal marker of mitosis progression and can be employed to differentiate vegetative (haploid) vs. planozygotes (diploid) cells. Thus, the FISH probe (oligo-Dino5Smix5) developed in this study facilitates analyses of the diversity, cell cycle and life stages of the genus Alexandrium.

鞭毛藻多样性和进化的染色体标记物很少，因为这些生物的基因组在真核生物中就其碱基组成和染色体结构而言是独特的。同样，缺少适当的工具也阻碍了对鞭毛藻无核小体染色体中5S核糖体DNA（rDNA）染色体定位的研究。在这项研究中，我们从亚历山大藻属的各种产毒素物种中分离并克隆了5S rDNA序列。并开发了荧光原位杂交（FISH）探针，可对其进行染色体定位。我们的结果可以概括如下：1）5S rDNA单元由一个高度保守的122 bp编码区和一个基因间隔区（IGS）组成，其长度和序列即使在菌株中也可变。2）在四个被研究物种（小菜曲霉，塔玛森曲霉，毛虫曲霉和太平洋曲霉）中发现了三种不同的IGS类型，其中一种包含U6小核RNA（snRNA）基因。3）在所有通过FISH调查的菌株中（小曲霉，塔玛生曲霉，太平洋曲霉，连叶曲霉，安德森曲霉和奥斯滕菲尔德曲霉），5S rDNA基因阵列与核仁组织区分开，核仁组织区包含45S前核糖体大RNA的基因。4）根据菌株/种类，每个单倍体基因组检测到1至3个5S rDNA位点。测定了A. minutum和A. pacificum菌株中5S rDNA位点的种内变异性。5）5S rDNA是有丝分裂进程的有用的染色体标记，可用于区分营养（单倍体）与扁平（二倍体）细胞。因此，在这项研究中开发的FISH探针（oligo-Dino5Smix5）有助于分析亚历山大藻属的多样性，细胞周期和生命阶段。