At the end of the 60-day experiment, 5 chickens were randomly selected from each treatment (a total of 4 treatments × 5 repeat = 20) and the cecal samples were collected in a cryopreservation tubes and then mixed and stored in liquid nitrogen at −70°C.
Microbial genome DNA was extracted from cecal samples by using QIAamp DNA stool mini kit (Tiangen Biotech, Beijing, China) according to the manufacturer’s instructions. PCR amplification of the V3–V4 region of the 16S ribosomal RNA (rRNA) gene was performed using the 341F/802R primer set (341F: CCTAYGGGRBGCASCAG; 802R: GGACTAC NNGGGTATCTAAT) as previously reported [17 (link)]. Only the products without primer dimers and contaminant bands were used for 16S rDNA high throughput sequencing at Shanghai Personal Biotechnology Co., Ltd (Shanghai, China).
Microbial genome DNA was extracted from cecal samples by using QIAamp DNA stool mini kit (Tiangen Biotech, Beijing, China) according to the manufacturer’s instructions. PCR amplification of the V3–V4 region of the 16S ribosomal RNA (rRNA) gene was performed using the 341F/802R primer set (341F: CCTAYGGGRBGCASCAG; 802R: GGACTAC NNGGGTATCTAAT) as previously reported [17 (link)]. Only the products without primer dimers and contaminant bands were used for 16S rDNA high throughput sequencing at Shanghai Personal Biotechnology Co., Ltd (Shanghai, China).
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