Sybr primescript rt pcr kit perfect real time

Total RNA was extracted with Trizol (TaKaRa) from shoots and roots. RNA was treated with RNase-free DNase (Transgen, China). First-strand cDNA was synthesized with the PrimeScript II 1st Strand cDNA Synthesis Kit (TaKaRa, Transgen, China). Quantitative real-time PCR was performed using the SYBR PrimeScript RT-PCR Kit (Perfect Real Time; TaKaRa). PCR was performed using a CFX 96 Real-Time system (Bio-Rad, Hercules, CA, United States) with the following standard cycling conditions: 95°C for 10 s, followed by 40 cycles of 95°C for 5 s, and 60°C for 30 s. Primer sequences used in the study was shown in Supplementary Table S1. The cycle threshold 2^(-ΔΔC(T))-based method was used for relative quantitation of gene expression. Expression levels of genes were normalized to Actin2.
Arabidopsis Genome Initiative locus identifiers for the genes mentioned in this article are as follows: ACTIN2 (AT3G18780), G6PD5 (AT3G27300), G6PD6 (AT5G40760), G6PD1 (AT5G35790), G6PD2 (AT5G13110), G6PD3 (AT1G24280), G6PD4 (AT1G09420), AtrbohD (AT5G47910), AtrbohF (AT1G64060), APX1 (At1G07890), SOD1 (At1G08830), POD1 (At1G67960), CAT1 (At1G20630), GR2 (At3G54660).

Total RNA was extracted with Trizol (TaKaRa) from roots, and then was treated with RNase-free DNase (Promega, Madison, WI, USA). First-strand cDNA was synthesized with the PrimeScript II 1st Strand cDNA Synthesis Kit (TaKaRa, Mountain View, CA, USA). Quantitative real-time PCR was performed using the SYBR PrimeScript RT-PCR Kit (Perfect Real Time; TaKaRa). PCR was performed using a CFX 96 Real-Time system (Bio-Rad, Hercules, CA, USA) with the following standard cycling conditions: 95 °C for 10 s, followed by 40 cycles of 95 °C for 5 s, and 60 °C for 30 s. The cycle threshold 2^{(−ΔΔC(T))}-based method was used for relative quantitation of gene expression. The specific primers for each gene are listed in Table S1. Expression levels of genes were normalized to ACTIN2 levels.