Identification of expression profiles of tapping panel dryness (TPD) associated genes from the latex of rubber tree (Hevea brasiliensis Muell. Arg.)

Abstract

Tapping panel dryness (TPD) occurrence in high latex yielding rubber tree (Hevea brasiliensis) is characterized by the partial or complete cessation of latex flow upon tapping leading to severe loss in natural rubber production around the world. The goal of this study was to identify genes whose mRNA transcript levels are differentially regulated in rubber tree during the onset of TPD. To isolate TPD responsive genes, two cDNA libraries (forward and reverse) from total RNA isolated from latex of healthy and TPD trees were constructed using suppression subtractive hybridization (SSH) method. In total, 1,079 EST clones were obtained from two cDNA libraries and screened by reverse Northern blot analysis. Screening results revealed that about 352 clones were differentially regulated and they were selected for sequencing. Based on the nucleotide sequence data, the putative functions of cDNA clones were predicted by BLASTX/BLASTN analysis. Among these, 64 were genes whose function had been previously identified while the remaining clones were genes with either unknown protein function or insignificant similarity to other protein/DNA/EST sequences in existing databases. RT-PCR analysis was carried out to validate the up-regulated genes from both the libraries. Among them, two genes were strongly down-regulated in TPD trees. The level of mRNA transcripts of these two genes was further examined by conventional Northern and RT-PCR analysis. Results indicated that the expression level of two genes was significantly lower in TPD trees compared to healthy trees. Many TPD associated genes were also up-regulated in TPD trees suggesting that they may be involved in triggering programmed cell death (PCD) during the onset of TPD syndrome. The results presented here demonstrate that SSH technique provides a powerful complementary approach for the identification of TPD related genes from rubber tree.

The Hevea brasiliensis AP2/ERF superfamily: from ethylene signalling to latex harvesting and physiological disease response

“…HbTCTP gene encodes cytosolic calcium binding protein that exact function remains to be elucidated in rubber tree. The transcript of this gene was also 60% lower during the occurrence of TPD in comparison to normal condition (Venkatachalam et al, 2007). In their study, Gébelin et al (2013) predicted that the microRNA, HbmiR159b could target the Myb-like transcription factor (HbMyb1) and glutathione peroxidase (HbGPX) in Hevea brasiliensis.…”

Histo- and cytopathology of trunk phloem necrosis, a form of rubber tree (Hevea brasiliensis Müll. Arg.) tapping panel dryness

“…Several structural abnormalities observed in the inner phloem of TPN-affected trees might also be related to the genes up-or downregulated in the latex or bark of TPD affected-trees (Chen et al 2002;Venkatachalam et al 2007Venkatachalam et al , 2009Li et al 2010). Notably, the disintegration of outer mitochondrial membranes in the trunk inner phloem of TPN-affected trees could be directly related to the HbTOM20 gene (Hevea brasiliensis Translocase of the Outer Mitochondrial Membrane) expression significantly downregulated in inner barks of TPD-affected trees compared with healthy one (Venkatachalam et al 2009).…”

Molecular cloning and characterization of phosphate (Pi) responsive genes in Gulf ryegrass (Lolium multiflorum L.): a Pi hyperaccumulator

“…However, limitations of global gene expression analysis due to the lack of entire genomic sequence information or microarrays for many of the model and crop species could be circumvented by suppression subtractive hybridization (SSH) technique. Differentially expressed genes could be normalized and enriched over 1,000-fold in a single round of hybridization by SSH and has been used successfully for identification of various biotic and abiotic stress responsive genes (Diatchenko et al 1999;Zhang et al 2005;Tian et al 2007;Venkatachalam et al 2007). …”