Development of a transformation system for Gracilaria changii ( Gracilariales , Rhodophyta ) , a Malaysian red alga via microparticle bombardment

Introduction Of the seaweed phycocolloids, agar has the higher price in the world market and is mostly extracted from red seaweeds. Critchley (1993) reported the world market value of agar as US $200 million. Hurtado-Ponce and Umezaki (1988) reported that the world's commercial agar comes mostly from gracilaroids. Agar extracted from Gracilariaceae became the first phycocolloid to be used in the human food industry (Armisen, 1995). Gracilaria has become an important agar source yielding 6070% of the world's agar (Schramm, 1991). Chile and China contribute 10,000 tonnes of agar annually. However, this cannot meet the demands for high quality agarose used extensively in biotechnology research. Malaysia has been importing agar from Korea, Japan, Hong Kong and China. In 1987, Malaysia imported 280 tonnes of agar worth more than RM 6.55 million (Jahara and Phang, 1990). In order for Malaysia to cope with the increasing demand for bacteriological agar in the coming years, there is a need to look into its own resources. In Malaysia, Gracilaria changii is one of the most abundant agarophytic seaweeds. Phang et. al. (1996) reported that Malaysian G. changii has potential commercial application in the agar industry due to its good quality agar and adaptability to the harsh mangrove conditions. Genetic engineering and manipulation will have an important role in the advancement of algal biotechnology based industries (Tseng, 1984; Van der Meer, 1988). The development of an efficient genetic transformation system will contribute to the genetic improvement of this commercially important seaweed. The basis for establishing a seaweed transformation system, is development of an efficient transformation method and selection of suitable reporter genes and promoters (Qin et al. 1998). Micro-particle bombardment and electroporation are commonly used in the transformation of marine microalgae and cyanobacteria (Matsunaga and Takeyama 1995; Toyomizu et al. 2001). However, microparticle bombardment is an effective and convenient method for transforming intact seaweed cells with cell walls (Qin et al. 1997, 1998, 1999). So far only a limited range of promoters have been employed in red seaweed transformation and only transient expression has been accomplished (Kubler et al. 1994). The most commonly used reporter gene is GUS gene (uid A). Transient expression of the lacZ reporter gene has only been achieved in Laminaria japonica (Qin et al. 1997), Undaria pinnatifida (Yu et al. 2002) and Haematococcus pluvialis (Teng et al. 2002) using SV40 promoter. Several studies on the application of SV40 promoter in obtaining stable transformation of macroalgae have been reported with L. japonica (Qin et al. 1997, 1998, 1999). Transgenic Laminaria with the hepatitis B surface antigen (HBsAg) gene has been obtained using SV40 promoter via particle bombardment (Jiang et al. 2002). In China, genetic and plant breeding has been applied in the cultivation industry of Laminaria japonica. The new varieties obtained yield 8-40% more biomass with 2058% higher iodine content than the control plants (Wu and Lin, 1987). With the genetic transformation system established for Gracilaria, genetic improvement can be carried out to optimise the exploitation of this commercial seaweed such as increase of biomass, improve agar/agarose quality, disease resistance, salt tolerance and other desire traits can be added to the seaweeds in culture. In future, there may be a possibility to use Gracilaria as bioreactors to produce vaccines, biochemicals and pharmaceuticals products.