2.6. RT-PCR Assay. RNA that was extracted using Trizol
reagent [19] frompassage five cellswas used as a template for
cDNA synthesis usingMoloneyMurine Leukemia Virus (MMLV,
Takara) according to the manufacturer’s instructions.
The primers were designed by primer 6.0 and were described
in Table 1. RT-PCR was continued for 35 cycles after an
initial denaturation at 94∘C for 10min. Each cycle of PCR
consisted of 94∘C for 30 sec, annealing temperature for 30 sec
and 72∘C for 30 sec, and a final extension for 10min at 72∘C.
PCR productions were visualized on a 2.5% agarose gel with
ethidium bromide [20].