红树林湿地生态系统中MCG古菌mcrA基因的时空变化规律和环境效应研究

The mcrA is the gene encoding to the α subunit of Methyl Co-enzyme M Reductase (MCR), the key protein involving in the methanogenesis. Because of the highly conserved sequence, mcrA gene is widely used as a molecular marker for diversity of methanogens, which was usually found in Euryarchaeota. Recently, high divergent mcrA genes were found in the genomes of newly proposed phylum, MCG archaea (Bathyarchaeota), suggest that mcrA genes have higher diversity than previously expected. However limit information has been documented for MCG mcrA gene. To investigate the diversity, abundance, expression, temporal and spatial distribution of mcrA gene from MCG archaea in mangrove sediments, we will collect mangrove sediment samples from two distinctive seasons at different sediment layers in different locations, using Illumina Hiseq high throughput sequencing, clone library, fluorescence in situ hybridization (FISH), quantitative PCR (qPCR) and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) technologies. Meanwhile, we will also analyze the relationships between environmental factors and the mcrA genes of MCG archaea in mangrove sediments to understand the responses of mcrA genes to environmental change, thus assessing their environmental effects. The results of this proposal will not only enhance the knowledge of mcrA gene, but also provide scientific data for understanding the methane metabolism of MCG archaea.

mcrA基因是微生物产甲烷途径的关键蛋白MCR（甲基辅酶M还原酶）的重要组成基因之一，是产甲烷古菌的分子标志物，主要发现于广古菌门。2015年Evan等在新的古菌门类MCG古菌（深古菌门）基因组中首次发现mcrA基因，其与已知mcrA有较大差别，提示该基因具有较高的多样性。然而，目前对MCG古菌中的mcrA基因还知之甚少。本课题拟以红树林湿地生态系统中的MCG古菌mcrA基因作为对象，在深圳红树林湿地设置不同的站位，采集不同季节和深度的沉积物样品，采用高通量测序、克隆文库、荧光原位杂交、定量PCR和qRT-PCR等技术，分析此基因的多样性、丰度及表达的时空变化规律；另外，结合环境参数，利用统计学方法分析MCG古菌mcrA基因与环境因子之间的内在关系，探讨其对于环境变化的响应并评估其环境效应。研究结果不仅可以丰富人们对mcrA基因的认识，同时也对评估MCG古菌在甲烷代谢中的作用提供理论支持。

mcrA基因是微生物产甲烷途径的关键蛋白MCR（甲基辅酶M还原酶）的重要组成基因之一，是产甲烷古菌的分子标志物，主要发现于广古菌门。2015年Evan等在新的古菌门类MCG古菌（深古菌门）基因组中首次发现mcrA基因，其与已知mcrA有较大差别，提示该基因具有较高的多样性。然而，目前对MCG古菌中的mcrA基因还知之甚少。本课题拟在深圳福田红树林湿地设置不同的站位，采集不同季节和深度的沉积物样品，采用高通量测序、克隆文库和定量PCR等技术，分析深圳福田红树林湿地中MCG古菌的分布和代谢多样性，以及mcrA基因尤其是MCG古菌的mcrA基因的丰度和分布。结果如下： . （1）深圳福田红树林湿地沉积物中古菌的主要种群包括了MCG古菌（Bathyarchaeota，深古菌）、奇古菌门（Thaumarchaeota）、广古菌门（Euryarchaeota）、洛基古菌门（Lokiarchaeota）和乌斯古菌门（Woesearchaeota）。其中MCG古菌随沉积物深度增加而逐渐增加，并且与总有机碳呈显著相关的关系。. （2）发现MCG古菌除了具有降解和利用蛋白质、多糖和芳香烃等多种有机物和生成乙酸的功能，还具备潜在的开尔文循环固碳、硫还原、氮还原产尿素、微生物B12产生和光敏感性等生理代谢途径。. （3）设计和评估了MCG古菌mcrA基因的引物，测定了深圳福田红树林湿地沉积物中的mcrA基因。结果表明深圳福田红树林湿地沉积物中的MCG古菌可能并没有携带mcrA基因。产甲烷八叠球菌目是深圳福田红树林湿地沉积物中最丰富的mcrA基因，其次是产甲烷微菌目和产甲烷杆菌目。并且mcrA基因的绝对丰度可能受不同季节中环境因子变化的影响。. 研究结果不仅丰富了人们对mcrA基因的认识，同时也对评估MCG古菌在甲烷代谢中的作用提供理论支持。.