3. Results
3.1. Construction of chimeric GLP-1 analogue/globulin protein expression vector We used a DNA encoding 30 amino acid residues corresponding to peptide [Ser8, Gln26, Asp34]-GLP-1 (7-36). The GLP-1 analogue gene was inserted into the variable region of the rice 26 kDa globulin gene between residues 109 and 110 (Fig. 1A) [18]. Furthermore, two junction sites of GLP-1 analogue and globulin were replaced by lysine and arginine residues, respectively, for trypsin digestion (Fig. 1A). In the present study, we used the new Hm/MAT-vector into which the hph gene and ipt gene [19,20] were inserted into the internal region between two directly oriented recombination sequences (RS) (Fig. 1B) [21]. The hph gene has been used as a selectable marker gene in addition to the ipt gene, and is eliminated from transgenic cells together with the recombines gene and the ipt gene. Therefore, the Hm/MAT-vector can generate selectable marker-free genetically modified rice seeds.
3. Results3.1. Construction of chimeric GLP-1 analogue/globulin protein expression vector We used a DNA encoding 30 amino acid residues corresponding to peptide [Ser8, Gln26, Asp34]-GLP-1 (7-36). The GLP-1 analogue gene was inserted into the variable region of the rice 26 kDa globulin gene between residues 109 and 110 (Fig. 1A) [18]. Furthermore, two junction sites of GLP-1 analogue and globulin were replaced by lysine and arginine residues, respectively, for trypsin digestion (Fig. 1A). In the present study, we used the new Hm/MAT-vector into which the hph gene and ipt gene [19,20] were inserted into the internal region between two directly oriented recombination sequences (RS) (Fig. 1B) [21]. The hph gene has been used as a selectable marker gene in addition to the ipt gene, and is eliminated from transgenic cells together with the recombines gene and the ipt gene. Therefore, the Hm/MAT-vector can generate selectable marker-free genetically modified rice seeds.
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