Expression of genes for E1 and CBH1 cellulase in transgenic tobacco and wheat
Creators: name: Inayath Hussain, Hashmath Hussain
Publisher: name: RMIT University
Year: 2024

description: Scarcity of fossil fuel coupled with ever-increasing requirements for energy in Australia and the rest of the world provide the impetus to look for alternative, sustainable energy sources. Production of alternative fuel, such as ethanol, from corn and sugar cane are today commercially viable operations in the USA and Brazil. The biomass used from corn and sugarcane is in the form of starch and sucrose, chemicals that are easily converted to ethanol. However, because corn and sugarcane are used for food, the fuel industry is competing with the food industry for these crops. Crop residues are produced in abundance in agriculture and forestry and are a potential source of plant biomass for ethanol production. Australia is a major wheat producer, and after harvest, there is a large biomass residue in the form of cellulose, rather than starch or sucrose. Like corn and sugar cane, cellulose from wheat, can be converted to glucose to generate ethanol but only after breakdown by cellulase enzymes. <br>There has been extensive research conducted into the bio-conversion of lignocellulose to soluble sugars. Presently, engineered organisms such as the yeast have been developed for simultaneous saccharification and fermentation process for producing novel cellulases required for converting complex sugar molecules to fermentable sugars (Mielenz, 2001). The process of producing enzymes by this method is expensive and alternative inexpensive means of enzyme production have been investigated (Austin-Phillips et al., 1999; Kawazu et al., 1999; Sakka et al., 2000). One approach is to express the cellulose enzymes in transgenic plants in the hope that the cost of enzyme production and hydrolysis of cellulose to fermentable sugars will be reduced. There are reports of successful cellulase expression in transgenic plants such as <em>Arabidopsis</em>, tobacco, duck weed, maize, and rice (Dai et al., 1999; Hood et at., 2007; Oraby et al., 2007; Sun et al., 2007; Ziegler et al., 2000), but more research is necessary to increase the level of cellulase enzyme expression in plants in order to make the process economical.<br>In this study plants were transformed with two genes; the Endoglucanase1 (E1) gene from <em>Acidothermu</em>. <em>cellulolyticus</em>, and the Cellobiohydrolase1 (<em>CBH1</em>) gene from <em>Trichoderma</em>. <em>reesei</em>. Genes for <em>E1</em> and <em>CBH1</em> were expressed in transgenic wheat plants. The expression of <em>E1</em>, accumulated up to 0.6% of the total soluble protein (TSP) in leaf material. More significantly, for the first time <em>CBH1</em> also accumulated to 0.25% of the TSP in leaf material, nearly twice the expression levels achieved in tobacco. The catalytic domain of E1 and CBH1 cellulases were also co-expressed for the first time in transgenic wheat. The recombinant cellulases expressed in wheat plants were observed to be biologically active. This work paves the way for the generation of transgenic wheat expressing genes for several different cellulases. These findings are significant given that wheat is, an important crop species and one of the largest sources of agricultural residue and potential sources of biomass for sustainable biofuel production.<br><br><br>