One of the major advantages of using DNA sequences to analyze microbiome diversity is that sequencing data obtained from different studies can be analyzed together, constituting a more cumulative approach than comparing DNA fingerprinting results [5]. However, if and how datasets from different sequencing projects can be combined for meta-analysis has not been evaluated because few studies have sequenced and compared actual microbiome

samples processed by different experimental methods. One of the most straightforward ideas is to use the same variable region for different PCR amplicons and extract this website sequences of that specific region from different studies for direct comparison. Theoretically, the same tag region allows for a consistent clustering of operational taxonomic units (OTUs) and taxonomy assignment; therefore, subsequent parameters, including α- and β-diversities selleck chemicals llc and community structures, can be analyzed. However, experimental conditions such as primer bias and sequencing quality might affect these analyses [11]. Until now, there have been no reports addressing this approach by amplifying real samples with different primers and extracting the same variable tag for direct comparison. In this study, we determined

a total of 28 fecal microbiome samples from four individuals and amplified each sample independently with two primer sets (V4F-V6R and V6F-V6R). We analyzed the α-diversity, β-diversity, microbial community structure, and biomarkers and focused on the following two questions: First, do the results from the two datasets agree with one another? Second, can the two datasets be combined

to produce reliable results? The present study provides useful information for evaluating the feasibility of meta-analysis for the study of microbiomes. Methods Ethical statement This study was approved by the Ethical Committee of Southern Medical University, and all participants provided written informed consent. Sample processing and sequencing Fecal samples were obtained from four individuals. For each individual, one sample was collected every two Etomidate days for a period of two weeks. All of the samples were stored at -80°C until DNA extraction, and 200 mg of each sample was used for DNA extraction. DNA was extracted using the PowerSoil DNA kit (MoBio, USA) according to the manufacturer’s instructions. The high fidelity ExTaq cocktail (Takara, China) was used to amplify the 16S rRNA gene tags. Each DNA sample was amplified by 2 barcoded primer sets, one of which included the primers V4F 5′ GTGCCAGCMGCCGCGGTAA 3′ and V6R 5′ ACAGCCATGCANCACCT 3′, while the other included the primers V6F 5′ CNACGCGAAGAACCTTANC 3′ and V6R 5′ ACAGCCATGCANCACCT 3′.