CHO表达生长激素释放因子的生物活性测定

Determination of the Activity of Growth Hormone Releasing Factor Expressed in CHO Cells

在对生长激素释放因子(GIF)基因改造和化学合成,并构建了其真核表达载体pCDNA3-GRF(1-32)的基础上,用Lipofectin将上述载体转染CHO细胞进行瞬时表达,采用体外单层垂体细胞培养的方法对表达的CRF(1-32)进行了体外生物活性测定,即先将垂体用酶进行消化,再将消化细胞培养,形成单层细胞,利用其能合成与释放生长激素的能力来检测GRF的活性。结果表明:表达产物可刺激生长激素释放,并且比对照组提高3.8倍。

Chemical synthesis of the GRF(1-32) gene had been finished in our former work. GRF(1032) gene, synthesized in this laboratory previously, was inserted into a plasmid , pcDNA3, to construct an expression vector in CHO cells. The expression vector was purified with a High Pure Plasmid Extraction Kit and then trasfected into prepared CHO cells in 60% confluent. Dispersed rat pituitary was used to determine the activity of expressed GRF(1-32) . The results showed that the expressed products stimulated Growth Hormone(GH) secretion in pituitary cells at high levels, being 3.8 times higher than the control group.