Molecular response of Hevea brasiliensis Ethylene Response Factors (HbERFs) as expression marker genes in response to ethephon stimulation in rubber tree clones

Real-Time quantitative RT-PCR technique is a sensitive method for measuring the accumulation of gene transcripts. This widely used technique in a variety of plant species; including rubber tree (Hevea brasiliensis) must meet basic criteria in order to produce accurate gene expression markers. Gene expression markers associated to the response of ethephon stimulation such as the Hevea brasiliensis Ethylene Response Factors (HbERFs) family has been characterized in a single rubber clone. It is known that the effect of genotype on rubber tree clones can give different expression of the same gene. This difference can be converted into a profile that characterizes clones to a certain trait. This study aimed to identify gene expression profile in response to ethephon stimulation using six HbERFs (HbORA47, HbRAP2.3, HbERF12, HbERF3, HbABR1, HbRRTF1) in three rubber tree clones having contrasted latex metabolism (PB 260, SP 217, and RRIM 600). Total RNA was isolated from 18 samples and used for cDNA synthesis. The quality of cDNAs was examined by PCR using HbActin primer. HbRH2b was selected among the 11 housekeeping genes to be used as an internal control in gene expression analysis. Gene expression analysis resulted to an induction and inhibition of HbERFs by ethephon stimulation which are specific to a particular clone. Expression profile of three Hevea clones showed distinct characteristics. The high latex metabolism clone PB 260 was characterized by the upregulated expression of HbRAP2.3 and HbERF12. The low latex metabolism clone SP 217 was characterized by the upregulated expression of HbRAP2.3 and HbRRTF1. Meanwhile, the profile of intermediate latex metabolism clone RRIM 600 was shown by downregulated expression of HbORA47 and up-regulated expression of HbABR1. This study shows that HbERFs gene family is an important expression marker because it can inform physiological conditions of rubber clones associated in response to ethephon.