Table 2.
Quantitative PCR validation of the CGH experiments
| Gene/region | forward primer | reverse primer | amplification efficiency (%) | reference strain | experimental strain | ratio (mean ± SD, n = 3) | ratio for region (CGH) |
|---|---|---|---|---|---|---|---|
| R26S (ref. gene) | GCTAGGCTTGCGTTTGTGTG | GGCGAGACAGAAAGATTCCG | 108 | strain 1 | strain 2/3 | - | - |
| Dynein (ref. gene) | GGAACAAAGCATGGTGACAACA | CGCGTGCCTATCCAAGCT | 97 | strain 1 | strain 2/3 | - | - |
| Sctg_16 | GCGTGCGTGCTTGGAAGG | TTCGGCTGCTGAGAGTGGAG | 96 | strain 1 | strain 3 | 0.9 ± 0.04 | 0.5 |
| Sctg_16 | CAACCGCTCTCCACCATTCAG | GACGCCTTCACAGTATCACACC | 96 | strain 1 | strain 3 | 0.7 ± 0.02 | |
| Sctg_16 | AACGATAGAGCGAGACGAGAGAG | GGAAGCAGATGGACACGAGTAAC | 93 | strain 1 | strain 3 | 0.8 ± 0.03 | |
| Sctg_68 | CTCCTATCGCCCTGTGGTCTC | ACTGCCTCTATGGTCCGTCTTG | 100 | strain 1 | strain 2 | 1.0 ± 0.1 | 0.4 |
| Sctg_68 | GTGAGAGAAACAACAGAGCAATACAG | ATGGAACCGCAGACAACAAGC | 102 | strain 1 | strain 2 | 0.6 ± 0.2 | |
| Sctg_68 | TCCGACCTGACGAGCATTGG | CAGTGTGCGGTGCGAACG | 103 | strain 1 | strain 2 | n/a* | |
| Sctg_68 | AAACACCTCCCAACCAACCAATC | AACGCAACGAGCAACCTTCC | 100 | strain 1 | strain 2 | n/a* | |
| EsV-1 | TAAGTTGATATTAGTGACAGTAGCAGGAG | GCCACGGAGGACGGAGATAC | 101 | strain 1 | strain 2 | n/a* | 0.5 |
| EsV-1 | ACCACGATGCCTGTCTCCTTAC | TCCTCAGCCGCCAGAATACG | 95 | strain 1 | strain 2 | n/a* | |
| EsV-1 | CTCCTCCGTAACCGTTGACATTG | CCGACCAGTAAACCCGTAAACC | 101 | strain 1 | strain 2 | n/a* | |
* no amplification in experimental strain, or difference > 10 cycles
Primers and validation results for each tested genomic region (EsV-1 = region on Sctg_52 containing the E. siliculous virus 1 genes, ref. gene = reference gene). The amplification efficiency was calculated from the standard curve for strain 1.