Ubiquitous anaerobic ammonium oxidation in inland waters Ubiquitous anaerobic ammonium oxidation in inland waters of China: an overlooked nitrous oxide mitigation process Guibing Zhu1,4*, Shanyun Wang1, Leiliu Zhou1, Yu Wang1, Siyan Zhao1, Chao Xia1, Weidong Wang1, Rong Zhou1, Chaoxu Wang1, Mike S. M. Jetten2, Mariet M. Hefting3, Chengqing Yin1, Jiuhui Qu1 1. Key Laboratory of Drinking Water Science and Technology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China 2. Department of Microbiology, Radboud University Nijmegen, the Netherlands; 3. Ecology and Biodiversity Group, Department of Biology, Utrecht University, the Netherlands; 4. Department of Biogeochemistry, Max Planck Institute for Marine Microbiology, Bremen, Germany * Corresponding author, E-mail:
[email protected] Supplementary Information Summary We provide here supplementary materials such as methodologies, complementary data, experimental analysis, supplementary figures and tables and detailed information of sampling sites. Detailed research method includes molecular (q)PCR, cloning and sequencing assay, measuring of anammox and denitrification rate with 15N-tracer technique by intact core method and slurry incubation method, N2O concentrations and fluxes measuring, and analytical procedures of environmental variables including physicochemical parameter and in-situ dissolved oxygen (DO). We also provided some supplementary figures and tables to illustrate the main text. The detailed information of sampling sites was listed in the last part. Ubiquitous anaerobic ammonium oxidation in inland waters Detailed research methods DNA Extraction, PCR, Cloning and Sequencing Analysis About 0.35 g freeze-dried sediment of each sample at each site was used for DNA extraction using a FastDNA SPIN Kit for Soil (Bio 101, USA) following the manufacturer’s protocol with some modifications.