Assessing effects of RT-PCR inhibitors in plant RNA extracts.

We conducted a mixing test in which we extracted nucleic acids (i) from leaf tissue of cultivated oat (Avena sativa cv. California Red; Poaceae; annual) infected with BYDV-PAV, from which a strong positive signal is easily recovered, and (ii) from this same infected oat tissue mixed 2:1 with uninfected switchgrass (Panicum virgatum; Poaceae; perennial) suspected to contain inhibitors. Two samples of each tissue type separately ground in liquid nitrogen were extracted by each of two commonly used extraction processes: (i) TRI Reagent from Molecular Research Center, Inc. (75 mg of tissue/sample), and (ii) a Spectrum Plant Total RNA kit from Sigma-Aldrich (100 mg of tissue/sample). We used a previously tested RT-PCR protocol to detect virus infection (55).