UNAIDS reported that 33.3 million people living with HIV at 2009. With current therapy, AIDS is still not curable or preventable. Griffithsin (GRFT) is a novel anti-HIV protein from the red alga Griffithsia that bind to HIV and inhibit cell to cell transmission. In this study first we optimized GRFT gene sequence based on codon frequencies for lettuce to reach highest expression in nuclear and chloroplast genome. In next step, we constructed pBI121 vectors containing GRFT gene with KDEL, extension and zera® signal peptides, a lettuce chloroplast transformation vector (PCL 96-39) and an expression vector (pET -28). For this purpose, first digestion with desired restriction enzymes was performed and after DNA purification from gel, ligation reactions and E. coli (Dh5α strain) transformation were carried out and resulted colonies used for confirmation of constructs. Digestion, PCR with gene specific and vector backbone primers as well as sequencing confirmed accurate and reliable integration of GRFT gene and signal peptides into corresponding vectors.
