Abstract
As a new expression system, Dunaliella salina (D. salina) has bright prospects and applications in various fields. However, its application is currently restricted because of the low expression and instability of foreign gene in D. salina cells. During genetic operation, transformation is a crucial step for genes expression in D. salina system. Although several transformation methods are existing currently, many inherent deficiencies and limitations still can be found in actual practice. Thus, we attempted to set up a rapid transformation method using the change of salt concentrations for D. salina. Based on osmotic pressure difference, exogenous genes can be spontaneously transferred into D. salina cells. After that, transformed D. salina cells were subjected to histochemical and molecular analysis. The results showed that the reporter gene, beta-glucuronidase genes were successfully expressed in the positive transformants, and detected in all of transformed cells by PCR analysis. Moreover, different transformation parameters, containing the salt gradient, time, dye dosage and Triton X-100 concentration, were optimized to obtain an optimal transformation result. Taken together, we preliminarily established a rapid transformation method with the features of fast, simple, economic, and high-efficient. This method will provide a strong genetic manipulation tool for the future transformation of D. salina system.
Highlights
Microalga as a versatile expression system that has been widely used in the fields of vaccine (Dehghani et al 2018), bio-based chemicals (Ng et al 2017), metabolic engineering (Anila et al 2016), pharmaceutical engineering (Zhang et al 2018), and so on
Morphology of stained D. salina cells with Ethidium bromide (EB) To measure the difference of treated D. salina lines, cells with fluorescence were counted under the fluorescence microscopy
To improve the gene expression in D. salina cells, and facilitate the maturation of D. salina system, a robust transformation tool is essential for production of recombinant proteins
Summary
Microalga as a versatile expression system that has been widely used in the fields of vaccine (Dehghani et al 2018), bio-based chemicals (Ng et al 2017), metabolic engineering (Anila et al 2016), pharmaceutical engineering (Zhang et al 2018), and so on. Since Dunaliella salina (D. salina) offers numerous special advantages, it has been exploited as a novel expression system for production of recombinant proteins (Poungpair et al 2014; Feng et al 2014a, b, c). Several exogenous genes have been transformed into D. salina, such as the human canstatin (Feng et al 2014a), soybean kunitz trypsin inhibitor (Chai et al 2012), and white spot syndrome virus VP28 (Feng et al 2014b), and interferon-thymosin fusion proteins (Zhang et al 2018), In the genetic operation of D. salina, transformation is a key step for expression of exogenous genes. More efficient and rapid approach should be established for future transformation of D. salina
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