Li Chunli, Han Lu, Zheng Zhenyu, Zhao Weidong
College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, China.
Sheng Wu Gong Cheng Xue Bao. 2011 Aug;27(8):1158-63.
According to the amino acid sequence of des-pGlu1-Brazzein, 4 pairs of oligonucleotide with cosmic site were synthesized by using yeasty biased codons. After linkage and PCR, the 179 bp code area of des-pGlu1-Brazzein was obtained and inserted into pPIC9K, which resulted in the recombinant expression vector pPIC9K-Bra. By digestion with Sal I, the lined pPIC9K-Bra was transformed into Pichia pastoris GS115 by electric shock. The results of expression indicted that the secreted target protein accounted for 51.6% of total protein in the supernatant and showed biological activity after purification.
根据去焦谷氨酸1-布拉泽因的氨基酸序列，利用酵母偏好密码子合成了4对含宇宙位点的寡核苷酸。经过连接和PCR，获得了去焦谷氨酸1-布拉泽因的179 bp编码区，并将其插入pPIC9K中，得到重组表达载体pPIC9K-Bra。用Sal I酶切后，将线性化的pPIC9K-Bra通过电击转化到毕赤酵母GS115中。表达结果表明，分泌的目标蛋白占上清液总蛋白的51.6%，纯化后具有生物活性。
