DNA sequence data collected during the SIPEX II voyage of the Aurora Australis, 2012

Purpose of experiments:Sequence data obtained to determine community structure of pack sea-ice microbial communities and whether it is effected by exposures to elevated CO2 levels.Summary of Methods:Cells in sea-ice brines were filtered onto 0.2 micron filters and material extracted using the MoBio Water DNA extraction kit. The DNA was analysed by Research and Testing Laboratories Inc. (Lubbock, Texas, USA) via 454 pyrosequencing. The bacteria were analysed using primers set 10F-519R, which targets 16S rRNA genes. 16S rRNA genes associated with chloroplast and mitochondria are included in this dataset but represent a minority of sequences in most samples. Eukaryotes were analysed using primers set 550F-1055R, which targets 18S rRNA genes. The 454 pyrosequencing analysis with the Titanium GS FLX+ kit used generates on average 3000 reads incorporating custom pyrotags for later stages of the data analysis. The specific steps used for subsequent data analysis are described in the attached PDF file (Data_Analysis_Methodology.PDF). This output was further refined by first determining consensus sequences at the 98% similarity level using Weizhong Li’s online software site CD-HIT (http://weizhongli-lab.org/cd-hit/) Reference: Niu B, Fu L, Sun S, Li W. 2010. Artificial and natural duplicates in pyrosequencing reads of metagenomic data. BMC Bioinformatics 1:187 doi:10.1186/1471-2105-11-187. The consensus sequences were then checked for errors, manually curated, and aligned against closest matching sequences obtained from the NCBI database (www.ncbi.nlm.nih.gov) to finally obtained a list of consensus operational taxonomic entities and the number of reads obtained for each samples analysed.File: SIPEXII_DNA_Sample_information.xlsx provides sampling and analysis information for the detailed results in the other two filesFile: SCIPEXII__sea_ice_bacteria_OTUs.xlsx contains information on the number of 16S rRNA reads in bacteria Phylum/Class and OTUsFile: SCIPEXII_sea_ice_brines_eukaryote_community_OTU_data.xlsx contains information on the number of 16S rRNA reads in eukaryotic microbes: Phylum/Order/Closest taxon and OTUs

实验目的：获取序列数据，以解析堆积海冰（pack sea-ice）微生物群落的群落结构，并探究其是否受高浓度CO₂暴露的影响。

方法概述：将海冰盐水（sea-ice brines）中的微生物细胞过滤至0.2微米孔径滤膜上，采用MoBio Water DNA提取试剂盒提取基因组DNA。DNA样本由美国德克萨斯州拉伯克市的研究与测试实验室有限公司（Research and Testing Laboratories Inc.）通过454焦磷酸测序进行分析。细菌群落分析采用靶向16S rRNA基因的引物对10F-519R。本数据集包含与叶绿体和线粒体相关的16S rRNA基因序列，但在多数样本中这类序列仅占少数。真核生物群落分析采用靶向18S rRNA基因的引物对550F-1055R。

本次实验采用Titanium GS FLX+测序试剂盒开展454焦磷酸测序，平均可产生3000条读段，并引入定制焦磷酸标签（pyrotags）用于后续数据分析环节。后续数据分析的具体步骤详见附件PDF文件（"Data_Analysis_Methodology.PDF"）。

首先借助Weizhong Li开发的在线软件CD-HIT（http://weizhongli-lab.org/cd-hit/），以98%的相似度阈值生成共识序列，对测序数据进行初步优化。相关参考文献：Niu B, Fu L, Sun S, Li W. 2010. 宏基因组数据焦磷酸测序读段中的人工与自然重复序列. BMC Bioinformatics 1:187 doi:10.1186/1471-2105-11-187。

随后对共识序列进行错误校验与人工整理，并与从美国国家生物技术信息中心（NCBI，www.ncbi.nlm.nih.gov）数据库获取的最匹配序列进行比对，最终得到共识操作分类单元（operational taxonomic unit, OTU）列表以及各分析样本对应的读段数量。

文件说明：
1. SIPEXII_DNA_Sample_information.xlsx 提供了另外两个文件中详细结果对应的采样与分析信息。
2. SCIPEXII__sea_ice_bacteria_OTUs.xlsx 包含细菌群落的16S rRNA读段数量信息，按门/纲及OTU进行分类。
3. SCIPEXII_sea_ice_brines_eukaryote_community_OTU_data.xlsx 包含真核微生物的16S rRNA读段数量信息，按门/目/最接近分类群及OTU进行分类。