Presently the green alga very attractive as a platform for various biotechnological and medical applications including its use as a biological factory for the production of high-value human therapeutic proteins. Agrobacterium mediated genetic transformation is a one kind of method for genetic transformation of the green alga, but there are few reports currently on the use of an Agrobacterium mediated transformation system. Just only limited species of microalgae were used and transformed via Agrobacterium to date. The development of an optimized and efficient Agrobacterium mediated transformation system is very important for microalgae to improve existing traits and enhance its economic potential.
In this present study, the establishment of an Agrobacterium-mediated transformation method and analysis of transgenic Chlorella vulgaris are demonstrated. After transformation with Agrobacterium tumefaciens strain LBA4404 harboring binary vector pCRFG, the chlorella vulgaris putative transgenic were regenerated on the selective medium containing 20 mg/L hygromycin and expression of the GUS activity. After 2 week, this transformed Chlorella showed to be GUS positive clones. For PCR analysis, using GUS gene and GFP gene specific primers that were designed to amplify the complete coding sequence of GUS and GFP. Amplification with GUS gene specific primers successfully detected the 947 bp and GFP gene specific primers successfully detected the 720 bp. GUS gene and GFP gene fragment from four putative transgenic lines. The results suggested that they were true Chlorella vulgaris transformants.
