Feng Xuehuan, Holzinger Andreas, Permann Charlotte, Anderson Dirk, Yin Yanbin
Department of Food Science and Technology, Nebraska Food for Health Center, University of Nebraska-Lincoln, Lincoln, NE, United States.
Department of Botany, University of Innsbruck, Innsbruck, Austria.
Front Plant Sci. 2021 Feb 10;12:610381. doi: 10.3389/fpls.2021.610381. eCollection 2021.
Zygnematophyceae green algae (ZGA) have been shown to be the closest relatives of land plants. Three nuclear genomes (, , and ) of ZGA have been recently published, and more genomes are underway. Here we analyzed two strains SAG 698-1a (mating +) and SAG 698-1b (mating -) and found distinct cell sizes and other morphological differences. The molecular identities of the two strains were further investigated by sequencing their 18S rRNA, and genes. These marker genes of SAG 698-1a were surprisingly much more similar to (SAG 698-2) than to SAG 698-1b. Phylogenies of these marker genes also showed that SAG 698-1a and SAG 698-1b were well separated into two different clades, where SAG 698-1a was clustered with , while SAG 698-1b was clustered with . Additionally, physiological parameters like ETR values differed between SAG 698-1a and SAG 698-1b after 2 months of cultivation. The de-epoxidation state (DEPS) of the xanthophyll cycle pigments also showed significant differences. Surprisingly, the two strains could not conjugate, and significantly differed in the thickness of the mucilage layer. Additionally, ZGA cell walls are highly enriched with sticky and acidic polysaccharides, and therefore the widely used plant nuclear extraction protocols do not work well in ZGA. Here, we also report a fast and simple method, by mechanical chopping, for efficient nuclear extraction in the two SAG strains. More importantly, the extracted nuclei were further used for nuclear genome size estimation of the two SAG strains by flow cytometry (FC). To confirm the FC result, we have also used other experimental methods for nuclear genome size estimation of the two strains. Interestingly, the two strains were found to have very distinct nuclear genome sizes (313.2 ± 2.0 Mb in SAG 698-1a vs. 63.5 ± 0.5 Mb in SAG 698-1b). Our multiple lines of evidence strongly indicate that SAG 698-1a possibly had been confused with SAG 698-2 prior to 2005, and most likely represents or a closely related species.
双星藻纲绿藻（ZGA）已被证明是陆地植物的近亲。最近已公布了ZGA的三个核基因组（ 、 和 ），更多基因组正在测序中。在这里，我们分析了两个ZGA菌株SAG 698 - 1a（可交配 +）和SAG 698 - 1b（可交配 -），发现它们的细胞大小和其他形态特征存在差异。通过对它们的18S rRNA、 和 基因进行测序，进一步研究了这两个菌株的分子特征。令人惊讶的是，SAG 698 - 1a的这些标记基因与 （SAG 698 - 2）的相似性远高于与SAG 698 - 1b的相似性。这些标记基因的系统发育分析也表明，SAG 698 - 1a和SAG 698 - 1b被很好地分为两个不同的 进化枝，其中SAG 698 - 1a与 聚类，而SAG 698 - 1b与 聚类。此外，培养2个月后，SAG 698 - 1a和SAG 698 - 1b的生理参数如ETR值有所不同。叶黄素循环色素的脱环氧化状态（DEPS）也显示出显著差异。令人惊讶的是，这两个菌株不能接合，并且在黏液层厚度上有显著差异。此外，ZGA细胞壁富含粘性和酸性多糖，因此广泛使用的植物核提取方案在ZGA中效果不佳。在这里，我们还报告了一种通过机械切碎进行高效核提取的快速简便方法，用于这两个SAG菌株。更重要的是，提取的细胞核进一步用于通过流式细胞术（FC）估计这两个SAG菌株的核基因组大小。为了确认FC结果，我们还使用了其他实验方法来估计这两个菌株的核基因组大小。有趣的是，发现这两个菌株的核基因组大小非常不同（SAG 698 - 1a为313.2 ± 2.0 Mb，而SAG 698 - 1b为63.5 ± 0.5 Mb）。我们的多条证据强烈表明，SAG 698 - 1a在2005年之前可能与SAG 698 - 2混淆，并且很可能代表 或与之密切相关的物种。
