Differences between single isolated cells and colonial cells in heteromorphic life cycle of Phaeocystis globosa：Morphology, Physiology and Transcriptome

purose:The large-scale reproduction of Phaeocystis globosa has caused serious damage to the marine ecosystem in the coastal waters of China. The outbreak of algae blooms depends on the competitive advantage of their heteromorphic life history: colonial formation is beneficial to resist zooplankton predation, and the single isolated cells can absorb nutrients rapidly. To better understand the different types of cells characteristics of P. globosa that facilitate rapid expansion, single isolated cells (GX-S) and colonial cells (GX-C) are separated by filtration. methods:For RNA exaction, the biomass was resuspended in 2 mL RNA extraction buffer (1:1 mix of aqua-phenol and buffer L [0.5% SDS, 10 mM EDTA, 0.2 M sodium acetate (pH 5), and 1:100 β-mercaptoethanol)] and then incubated with DNase I (Takara, Japan) for 30 min at 37 °C to remove genomic DNA. RNA quality analysis, library construction, sequencing, data filtering and mapping were performed by the Novogene Bioinformatics Technology Co., Ltd. (Beijing, China). mRNA profiles of 10 days GX-C and GX-S were generated by deep sequencing, in triplicate. results:Three biological replicates from GX-S and GX-C were ensured statistical comparability and reliability of data. Raw data ranged from 25, 608,632 to 35,516,726 reads per sample. After producing more than 24 million clean reads, removing low-quality sequences and adapter sequences. Additionally, 4386 genes were differentially expressed at statistically significant levels, which included 2268 up-regulated genes and 2118 down-regulated genes. Genes with significant differential expression were involved in several pathways, including starch and sucrose, phagosome, inositol phosphate metabolism, fatty acid degradation. conclusions: In summary, we can find that colonial cells have stronger carbon fixation capacity. It is not used to synthesize fatty acids as reserves of energy, but to secrete EPS. The reduction of fatty acid makes P. globosa become “low-quality food” and the formation of colony from EPS reduces the chance of being ingestion by zooplankton. mRNA profiles of 10 days GX-C and GX-S were generated by deep sequencing, in triplicate, using Illumina HiSeq

研究背景：球形棕囊藻（Phaeocystis globosa）的大规模增殖已对中国近海水域的海洋生态系统造成严重破坏。赤潮暴发依赖于其异形生活史的竞争优势：群体形成有助于抵御浮游动物捕食，而单个游离细胞可快速吸收营养物质。为更好地理解球形棕囊藻不同细胞类型促进其快速扩张的特性，研究人员通过过滤法分离得到单个游离细胞（GX-S）与群体细胞（GX-C）。
实验方法：RNA提取时，将生物样本重悬于2 mL RNA提取缓冲液（水饱和苯酚与L缓冲液按1:1体积比混合，L缓冲液成分为0.5%十二烷基硫酸钠（SDS）、10 mM乙二胺四乙酸（EDTA）、0.2 M乙酸钠（pH 5），并添加1:100体积的β-巯基乙醇），随后于37 ℃下与脱氧核糖核酸酶I（DNase I，宝生物工程（Takara），日本）孵育30分钟以去除基因组DNA。RNA质量检测、文库构建、测序、数据过滤与比对工作均由北京诺禾致源生物信息科技有限公司（Novogene Bioinformatics Technology Co., Ltd.，中国北京）完成。本研究对培养10天的GX-C与GX-S进行了三次生物学重复的深度测序，以获取其mRNA表达谱。
实验结果：本研究确保GX-S与GX-C的三个生物学重复具备统计学可比性与数据可靠性。每个样本的原始读段数量介于25,608,632至35,516,726之间。在获得超过2400万条清洁读段后，过滤掉了低质量序列与接头序列。此外，共鉴定得到4386个具有统计学显著性差异的表达基因，其中2268个基因上调表达，2118个基因下调表达。显著差异表达基因参与了多条代谢通路，包括淀粉与蔗糖代谢、吞噬体通路、肌醇磷酸代谢以及脂肪酸降解通路。
研究结论：综上可见，群体细胞具备更强的固碳能力。其并不会将碳源用于合成脂肪酸作为能量储备，而是用于分泌胞外多糖（EPS）。脂肪酸含量的降低使得球形棕囊藻成为“低质量食物”，而由胞外多糖形成的藻落则降低了被浮游动物摄食的概率。本研究通过Illumina HiSeq平台，对培养10天的GX-C与GX-S进行了三次生物学重复的深度测序，获取了其mRNA表达谱。