| Natural rubber, which is produced as a secondary plant metabolite, has been well recognized as an important raw industrial material due to its unique properties that can not be substituted by any alternative synthetic ones. This makes the worldwide supply of natural rubber is barely keeping up with its demand, and such resultant gap is expected to be wider even more significantly in the future. Although more than 2000 plant species have been shown to possess the ability to synthesize natural rubber, Hevea brasiliensis is yet the only one qualified as a commercial source for natural rubber production. The large yield, high quality, and easy manipulation of culture and harvest all count Hevea brasiliensis as a predominant natural rubber provider with valuable economical significance. However, the fact that the limited rubber supply due to the limited rubber plantation is far behind the ever-stringent demand makes it necessitated for exploration of new approaches aimed for significant increase of the yield of Hevea brasiliensis.Advances by employing biotechnological principles in rubber tree bioengineering process emerges in a much slower pace compared to the studies by use of model crops. The putative cDNA candidates so far isolated from Hevea Brasiliensis, rather than being originated directly as novel genes, were most obtained through homologous cloning based on findings derived from other specifies. Moreover, the mechanism(s) of latex regeneration remains largely unknown. The focusing points currently under intensive studies is delineation ofrubber transferase gene and the regulatory networks engaged in dynamic control of this gene expression, since rubber transferase is the key enzyme fundamental to rubber biosynthesis.The notion that the capacity of laticiferous system of a rubber tree is the main limiting factor in the control of the latex regeneration between the tappings makes it essential for identification and characterization of genes relevant to latex regeneration in laticiferous system. Furthermore, findings fruitful from these studies will greatly benefit the bio-breeding of high-yield strains. In addition, the perception of laticiferous system of Hevea brasiliensis as an ideal bioreactor for large-scale production of heterologous bioactive substances (e.g. PHB, poly-3-hydroxybutyrate), insights into the molecular mechanism(s) of latex regeneration will no doubt accelerate the progress in this field.The current investigation is objective to identify and characterize novel genes expressed in latex of Hevea Brasiliensis The studies were initiated by construction of a subtraction cDNA library by employing suppressive subtractive hybridization (SSH) methodology, which were followed by intensive systematic analyses of full-length cDNA clones or cDNA fragments that are only positively correlated with latex regeneration. Summery of the results: 1. Construction and screening of a substraction cDNA libraryA subtracted cDNA library was constructed via suppressive subtractive hybridization, in which the latex poly(A+)RNA isolated from regularly tapped trees was utilized as tester while driver is poly(A+)RNA derived from negative latex-producing leaf tissues. The strategy, therefore, maintain tester and driver to harbor identical genetic background, which maximum eliminates potential false positives that are very likely incurred by use of materials with non-equivalent origins. Moreover, due to the simultaneous subtraction of non-differentially expressed genes in tester, genes particularly associated with laxtex-regeneration stages were specifically enriched.Total 79 positive clones obtained from screening of the subtract library constructed were further verified by analyses using reverse Northern dot blots. 18 out of these 79 confirmed clones were randomly selected for sequencing analyses, of which the results were aligned with GenBank?databases via BLASTn and BLASTx programs. It is shown that 7 of 18 candidates have been indicated to exert a positive role in rubber synthesis. Six of them are a... |
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