Table 2.
16S rRNA-targeted primers sequences, concentrations, and qPCR conditions used in this work for SYBR green chemistry.
| Target* | Primers |
Concentration
(nM) |
Sequence (5′-3′) | Reference | qPCR conditions | |||||
|---|---|---|---|---|---|---|---|---|---|---|
| Denaturing | Annealing (40 cycles) | Melting curve | ||||||||
| Time (min:s) | T a (°C) | Time (min:s) | T a (°C) | Time (min:s) | T a (°C) | |||||
| EUB | Eub_F Eub_R |
200 | ACTCCTACGGGAGGCAGCAGT GTATTACCGCGGCTGCTGGCAC |
(Matsuda et al., 2011)¶ | 10:00 | 95 | 0:15 1:00 |
95 54 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
| AKK | AKK_F AKK_R |
250 | CAGCACGTGAAGGTGGGGAC CCTTGCGGTTGGCTTCAGAT |
(Sekirov et al., 2010) | 10:00 | 95 | 0:15 1:00 |
95 60 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
| ROS | ROS_F ROS_R |
150 | TACTGCATTGGAAACTGTCG CGGCACCGAAGAGCAAT |
(Larsen et al., 2010) | 10:00 | 95 | 0:15 1:00 |
95 60 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
| RUM | RUM_F RUM_R |
250 | GGCGGCYTRCTGGGCTTT CCAGGTGGATWACTTATTGTGTTAA |
(Round and Mazmanian, 2009) | 10:00 | 95 | 0:15 1:00 |
95 60 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
| LAC | VLAC_F VLAC_R |
200 | AGCAGTAGGGAATCTTCCA CGCCACTGGTGTTCYTCCATATA |
(Payne et al., 2012) | 10:00 | 95 | 0:15 1:00 |
95 60 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
| FIR | FIR_F FIR_R |
150 | TGAAACTYAAAGGAATTGACG ACCATGCACCACCTGTC |
(Mühling et al., 2008) | 10:00 | 95 | 0:15 1:00 |
95 60 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
| BAC | BAC_F BAC_R |
300 | CRAACAGGATTAGATACCCT GGTAAGGTTCCTCGCGTAT |
(Lozupone et al., 2012) | 10:00 | 95 | 0:15 1:00 |
95 60 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
| XIV | VXIV_F VXIV_R |
300 | CGGTACCTGACTAAGAAGC AGTTTYATTCTTGCGAACG |
(Ramirez-Farias et al., 2009) | 10:00 | 95 | 0:15 1:00 |
95 60 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
| B46 | B46_F B46_R |
300 | TCCACGTAAGTCACAAGCG CGCCTACCTGTGCACTACTC |
(Malagón et al., 2019) | 10:00 | 95 | 0:15 0:45 |
95 62 |
1:00 0.30 |
95 55 |
| 0:30 | 95 | |||||||||
*
AKK, Akkermansia muciniphila; ROS, Roseburia species; RUM, Ruminococcus species; LAC, Lactobacillus; FIR, Firmicutes; BAC, Bacteroidetes; XIV, Clostridial cluster XIV; B46, S. variabile.
¶
Modified from.