16S ribosomal ribonucleic acid sequencing

YI Yumiko Ikubo
TS Takayuki Jujo Sanada
KH Koji Hosomi
JP Jonguk Park
AN Akira Naito
HS Hiroki Shoji
TM Tomoko Misawa
RS Rika Suda
AS Ayumi Sekine
TS Toshihiko Sugiura
AS Ayako Shigeta
HN Hinako Nanri
SS Seiichiro Sakao
NT Nobuhiro Tanabe
KM Kenji Mizuguchi
JK Jun Kunisawa
TS Takuji Suzuki
KT Koichiro Tatsumi
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Amplification and sequencing of bacterial 16S rRNA was performed according to our previous reports [25]. The V3-V4 region of the 16S rRNA gene of bacteria was amplified by polymerase chain reaction (PCR) using the following primers: forward, 5′-TCGTCGGCAGCGTCAGATGTGTATAAGCGACAGCCTACGGGNGGCWGCAG-3′, and reverse, 5′-GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGGACTACHVGGGTATCTAATCC-3′, which was followed by the addition of the sequencing adapters. The amplicons were sequenced using the Illumina MiSeq platform (Illumina Inc., San Diego, CA, USA). A total of 10,000 reads per sample were randomly selected for further analysis. Samples with insufficient read numbers were re-sequenced, and those with repeated insufficient read numbers were excluded.

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