By Alfred Haas
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Reverse: Tilletia-R (5′-CAA GAG ATC CGT TGT CAA AAG TTG-3′). 2. Thaw the working solutions from the stock solutions and make up the reaction mix (as shown below in Table 5) for the appropriate number of reactions. 2 mL PCR tubes. Add the template DNA to the individual PCR tubes containing the reaction mix. NB: If using the single spore from the previous step (DNA extraction step), add 19 μL of the reaction mix directly to this Eppendorf tube and crush the coverslip sandwich using the pipette tip.
In potato 19 However, at the molecular level there is considerable similarity between these two closely related Phoma varieties. MacDonald et al.  developed a RAPD-generated PCR-RFLP marker to distinguish between the two varieties, but the use of restriction enzymes makes this assay time-consuming and relatively expensive. Cullen et al.  developed conventional and quantitative PCR assays for the detection of P. exigua var. foveata, but the primers also detected the closely related P. exigua var.
The multiplex assay achieves the same level of sensitivity as the individual singleplex reactions. The multiplex detection limit is 80 fg for P. foveata (Fig. 2b), 160 fg for Phoma species, and 80 fg for P. eupyrena. 98 for P. foveata, Phoma complex, and P. eupyrena, respectively, indicating linear responses in detection related to the increasing DNA concentration, which can be used to estimate DNA concentration of unknown samples. Figure 2c shows the standard curve obtained for Phoma complex amplification of P.
Oscilloscope techniques by Alfred Haas