Figure 3
Optimization of duplex PCR. Alleles were amplified with a duplex set of SSR primer pairs (PVCAG-2517/8 and PVCAG-2397/8). Eight switchgrass genotypes from Alamo and Kanlow were used as amplification templates. The adjustments of PCR reaction components were shown from group A to H. Group A: control PCR with the same conditions as monoplex PCR, except for mixing PVCAG-2517/8 and PVCAG-2397/8 together; B: doubling Taq polymerase; C: doubling dNTPs concentration; D: doubling concentrations of DNA templates; E: increasing buffer concentration to 1.6x; F: increasing Mg 2+ concentration to 2.4 mM; G: doubling IR-M13 dye; H: doubling primer concentrations of PVCAG-2517/8 and PVCAG-2397/8. “M” indicated the DNA ladder.