Dice real time system tp800

Total RNA extracted from N. benthamina leaves or protoplasts were subjected to reverse transcription using PrimeScript RT reagent Kit (Takara) using oligo-dT and random primers according to manufacturer’s protocol. Real-time PCR was carried out using SYBR Premix Ex Taq (RR420A, Takara). Primers used for real-time PCR analysis were listed in S2 Table. Quantitative analysis of each mRNA was performed using a Thermal cycler Dice Real Time System TP800 (Takara).

Total RNA was extracted from conditioned seeds before GR24-treatment, purified with the Total RNA Extraction Mini Kit (RBC Bioscience), and converted to cDNA with the PrimeScript RT Reagent Kit (Takara Bio) according to the manufacturer’s protocols. Quantitative PCR was performed with SYBR Premix Ex Taq (Takara Bio) and the Thermal Cycler Dice Real Time System TP800 (Takara Bio). The transcript levels of ShKAI2iB were normalized against those of ShUBQ145 (link), using primers specific for ShKAI2iB (5′-TAGGGTCGGTGGAAGGTCAGTC-3′ and 5′-CAGCACTGGGATGGCAACCT-3′), and ShUBQ1 (5′-CATCCAGAAAGAGTCGACTTTG-3′ and 5′-CATAACATTTGCGGCAAATCA-3′). Student’s t-test was used to determine the significance of differences relative to the transcript level in Striga seeds conditioned for 1 day.

Total RNA extracted from N. benthamina leaves or protoplasts were subjected to reverse transcription using PrimeScript RT reagent Kit (Takara) using oligo-dT according to manufacturer's protocol. Real-time PCR was carried out using SYBR Premix Ex Taq (RR420A, Takara) using primers 29 and 30 for EF1 and primers 16 and 31 for NbGAPDH-A. Quantitative analysis of each mRNA was performed using a Thermal cycler Dice Real Time System TP800 (Takara). Semi-quantitative RT-PCR was performed using the same cDNA and primers and amplified by Ex Taq polymerase (Takara).