Each collected swab was randomly allotted to a new identifier for blindness. Bacterial DNA was extracted from all samples using the Qiagen DNA Extraction kit (Qiagen, Hilden, Germany) according to the modified protocol developed by the laboratory of Gary B Huffnagle.13 (link) Amplification and sequencing of throat samples were performed at BGI (Shenzhen, China) in terms of Roche 454-based sequencing protocols.14 (link),15 (link) Briefly, the V3–V5 regions of the gene encoding 16S ribosomal RNA (rDNA) were amplified by polymerase chain reaction using the forward primer (5′-CCGTCAATTCMTTTGAGTTT-3′) and the reverse primer (5′-ACTCCTACGGGAGGCAGCAG-3′) with sample-specific barcodes. After purification of amplicons, sequencing was conducted through 454 platform (Roche Applied Science, Basel, Switzerland).
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Bacterial DNA Profiling of Throat Microbiome
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Other organizations : Peking University Third Hospital, Peking University
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- Randomized allotment of collected swabs to new identifiers for blindness
- Bacterial DNA extracted from all samples
- Amplification and sequencing of throat samples
- Use of Qiagen DNA Extraction kit (Qiagen, Hilden, Germany) according to the modified protocol developed by the laboratory of Gary B Huffnagle
- Amplification and sequencing of the V3–V5 regions of the gene encoding 16S ribosomal RNA (rDNA) using the forward primer (5′-CCGTCAATTCMTTTGAGTTT-3′) and the reverse primer (5′-ACTCCTACGGGAGGCAGCAG-3′) with sample-specific barcodes
- Sequencing conducted through 454 platform (Roche Applied Science, Basel, Switzerland)
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