Computational protocol: Changes in land use driven by urbanization impact nitrogen cycling and the microbial community composition in soils

Similar protocols

Protocol publication

[…] The V4 and V5 regions of the 16 S rRNA gene were amplified with 515 F/907 R primers. The reverse primers (907 R) were tagged with a six-base barcode (). The procedures to process the sequencing were displayed in ().Sequence data were analyzed using QIIME version 1.9.1. Raw sequences were demultiplexed and low quality or ambiguous reads were removed and then chimeric reads were removed and checked. Filtered sequences were clustered into operational taxonomic units (OTUs) at 97% similarity level and the representative sequence for each OTU was selected. The representative sequences were then assigned to taxonomy using RDP. OTUs with a single sequence or assigned to mitochondria or chloroplast were filtered. Samples were then rarefied to determine the alpha diversity and beta diversity (). LEfSe was calculated to identify the corresponding taxa with higher abundance in different samples ().All the sequences were submitted to the European Nucleotide Archive of EMBL. The accession number is PRJEB14752. [...] Considering the unbalanced sample size in three categories, the nonparametric Kruskal-Wallis H test was employed to observe the significance of urbanization impact on the potential activities and gene abundances. To compare the significance of variation between each two of categories, post hoc multiple comparisons were conducted using the Dunnett’s T3 method. Fisher’s Least Significant Difference (LSD) was calculated as previously described, to compare the significance between each two sites. The counts in OTU table were transformed to relative abundances as percentage. CCA was then performed to determine the significant factors shaping the bacterial communities. Variation partitioning was used to reveal the proportions of variance explained by different factors by running the partial CCA (). A heatmap was generated to exhibit the relative abundance of phylum taxa in different samples. Pearson’s correlation analysis was conducted between each pair of the parameters or values measured in this study. ANOSIM and PERMANOVA were employed to test the significance of urbanization (urban, suburban and rural) impact on the beta-diversity based on the Unifrac matrices. Mantel tests were performed to explore the correlations between different variables and microbial communities based on the Unifrac matrices. Kruskal-Wallis H test, multiple comparisons and correlation analysis were carried out with SPSS v19.0 software (IBM, USA). The other analyses were performed using R v3.3.2 software (R Foundation for Statistical Computing, Vienna, Austria) (). The null hypothesis was rejected when p-values were less than 0.05. […]

Pipeline specifications