TABLE 2.
Oligonucleotide sequences developed for assay of B. abortus
| Oligonucleotide | Position and orientation (B. abortus numbering; gb AF148682) | Sequence |
Tm (°C)a
|
Reference | ||
|---|---|---|---|---|---|---|
| TM Utility, version 1.3 | MeltCalc 2.06 Pro | R.A.P.I.D. Exp. | ||||
| BAF primer | Forward (1381-1399) | 5′-CCATTGAAGTCTGGCGAGC-3′ | 67.6 | 68.1 | 86.5 (amplicon) | This work |
| BAR primer | Reverse (1537-1516) | 5′-CGATGCGAGAAAACATTGACCG-3′ | 68.7 | 69.4 | This work | |
| TH 5′-exonuclease probe | Reverse (1492-1473) | 5′-FAM-GCATGCGCTATGATCTGGTTACGTT-(TAMRA)-3′ | 70.4 | 70.7 | Hadfield (unpublished) | |
| BAFR FAM hybridization probe | Reverse (1470-1489) | 5′-TGCGCTATGATCTGGTTACG-(FAM)-3′ | 64.9 | 64.6 | 63 (probe) | This work |
| BAFR CY5.5 Hybridization probe | Reverse (1467-1450) | 5′-(CY5.5)-AAATGCAGACACGCCCTA-(P)-3′ | 65.7 | 66.1 | This work | |
a
The Tm for the primer or probe was calculated by using Idaho Technology's TM Utility version 1.3 with optimized conditions for each reaction, MeltCalc 2.06 Pro, or R.A.P.I.D. For R.A.P.I.D., the values shown represent the average experimental value based on R.A.P.I.D. melt curve analysis with B. abortus NADC 1144 as the template.