Reference Gene Selection For Quantitative Real-time PCR In Maize Treated With Abiotic Stresses And Hormones

Maize is one of the most important food crops, animal feed and cash crop in our country. However, with the increasing deterioration of the environment, the abiotic stress become more and more serious, which affect the quality and output of crop. Phytohormones also play an important role in regulating the plants response to the abiotic stress and improving the adaption to abiotic stress. Therefore, studys about plants abiotic stress and phytohormones signalling related to the abiotic stress are highlighted all the time in the research field of botany. Gene functional analysis is a prerequisite for the utilization of this gene in providing favorable safeguard and reducing the impact caused by abiotic stress. Gene expression pattern reflects to some extent of the gene function. At present, RT-qPCR tends to be used to detect gene expression due to the special advantages. In order to ensure the reliability of the RT-qPCR results, appropriate internal reference genes should be choosed. However, there is no gene could express steadily across various conditions such as different treatments, tissues and developmental stages. Therefore, an appropriate assessment of the expression stability of candidate reference genes is necessary before its use for normalization of expression data.In this study, the expression of eight widely used internal control genes(GAPDH, EF1α, TUB, ACT2, UBQ9, CYP, EIF4A and UBQ7) together with two genes (GLU1and RBA15) selected from the microarray datasets were detected in maize seedling under different abiotic stress (cold stress, heat stress, drought stress, salt stress and Al toxicity) and hormone treatment (SA, JA, ABA, GA and ACC) by RT-qPCR. Afterwards, the expression stability of these genes was analysed by geNorm, NormFinder and BestKeeper programs and finally the most appropriate reference genes and their numbers were analyzed. The results were as fellows:1. Under abiotic stress conditions:CYP, EIF4A and EF1α were the most appropriate reference genes under cold stress; EF1α, ACT2, CYP and EIF4A were the most appropriate reference genes under heat stress; TUB and GAPDH were the most appropriate reference genes under drought stress; EF1α, TUB, GAPDH, CYP, EIF4A and ACT2were the most appropriate reference genes under salt stress; CYP and EIF4A were the most appropriate reference genes under Al toxicity.2. Under hormone treatment:EF1α, ACT2and UBQ9were the most appropriate reference genes in SA treatment; TUB and GAPDH were the most appropriate reference genes in JA treatment; EF1α, TUB and GAPDH were the most appropriate reference genes in ABA treatment;ACT2and TUB were the most appropriate reference genes in GA treatment; EF1α and TUB were the most appropriate reference genes in ethylene treatment.3. In different tissues under different treatment:EF1α, UBQ9, TUB and CYP were the most appropriate reference genes in maize roots; TUB, UBQ9and EF1α were the most appropriate reference genes in maize stems; EF1α, UBQ9and GAPDH were the most appropriate reference genes in maize leaves.