Semi-quantitative RT-PCR analysis of transcripts encoding protease inhibitor in Hevea brasiliensis Muell. Arg latex

Abstract

In this paper, the semi-quantitative RT-PCR was carried out on four Hevea brasiliensis protease inhibitor transcripts (HbPI1-4) of rubber tree clone PB 260 under abiotic and biotic stresses. The research aimed to understand whether the expressions of HbPIs are induced by either biotic or abiotic stresses. The RNA isolation was performed under strict procedure to synthesize the best quality of cDNA. The accumulation of gene transcript was determined by RT-PCR (Reverse Transcriptase – Polymerase Chain Reaction) at the 30th cycle. Pre-PCR experiments were carried out on 20^th, 25^th, 30^th, and 35^th cycles for each gene. At the 30^th cycle, PCR products were clearly visible on an agarose gel and could be quantified in comparison to the reference gene (HbActin). In general, the value of gene expression of HbPIs ranged from 0.222 to 0.531 in various conditions. However, the data were not statistically different under abiotic (tapping panel dryness, TPD) and biotic (white root disease, WRD) stresses. The semi-quantitative RT-PCR was able to confirm the relatively stable expression of HbPIs under five tree conditions (healthy trees, TPD-affected trees at <50% dry cut length (DCL), TPD-affected trees at >50% DCL, WRD-affected trees and both of symptoms).