Zhang Zehao, Wang Junhao, Zhang Xiaoqian, Guan Xiaowei, Gao Tian, Mao Yunxiang, Poetsch Ansgar, Wang Dongmei
Key Laboratory of Marine Genetics and Breeding (OUC), Ministry of Education, Qingdao 266000, China.
College of Marine Life Sciences, Ocean University of China, Qingdao 266000, China.
Plants (Basel). 2023 May 5;12(9):1883. doi: 10.3390/plants12091883.
Contamination from cytosolic DNA (plastid and mitochondrion) and epiphytic bacteria is challenging the efficiency and accuracy of genome-wide analysis of nori-producing marine seaweed . Unlike bacteria and organellar DNA, nuclear DNA is closely associated with histone proteins. In this study, we applied Chromatin Immunoprecipitation (ChIP) of histone H3 to isolate nuclear DNA, followed by high-throughput sequencing. More than 99.41% of ChIP-sequencing data were successfully aligned to the reference nuclear genome; this was remarkably higher than those from direct extraction and direct extraction data, in which 40.96% to 42.95% are from plastids. The proportion of data that were mapped to the bacterial database when using ChIP extraction was very low. Additionally, ChIP data can cover up to 89.00% of the nuclear genome, higher than direct extraction data at equal data size and comparable to the latter at equal sequencing depth. The uncovered regions from the three methods are mostly overlapping, suggesting that incomplete sequencing accounts for the missing data, rather than failed chromatin-antibody binding in the ChIP extraction method. This ChIP extraction method can successfully separate nuclear DNA from cytosolic DNA and bacterial DNA, thus overwhelmingly reducing the sequencing cost in a genome resequencing project and providing strictly purified reference data for genome assembly. The method's applicability to other macroalgae makes it a valuable contribution to the algal research community.
来自胞质DNA（质体和线粒体）以及附生细菌的污染，正挑战着紫菜生产海洋藻类全基因组分析的效率和准确性。与细菌和细胞器DNA不同，核DNA与组蛋白紧密相连。在本研究中，我们应用组蛋白H3的染色质免疫沉淀（ChIP）来分离核DNA，随后进行高通量测序。超过99.41%的ChIP测序数据成功比对到参考核基因组；这显著高于直接提取数据，直接提取数据中40.96%至42.95%来自质体。使用ChIP提取时，比对到细菌数据库的数据比例非常低。此外，ChIP数据可覆盖高达89.00%的核基因组，在相同数据量时高于直接提取数据，在相同测序深度时与后者相当。三种方法未覆盖的区域大多重叠，这表明测序不完整导致了数据缺失，而非ChIP提取方法中染色质 - 抗体结合失败。这种ChIP提取方法能够成功地将核DNA与胞质DNA和细菌DNA分离，从而极大地降低了基因组重测序项目中的测序成本，并为基因组组装提供了严格纯化的参考数据。该方法对其他大型藻类的适用性，使其对藻类研究界做出了有价值的贡献。
