人凝血因子IX在脆壁克鲁维酵母中的表达与分泌

通过改造人凝血因子IX基因(h-FIX)使其与乳酸克鲁维酵母(Kluyveromyces lactis)的分泌信号序列(Kss)融合，并置于脆壁克鲁维酵母(K．fragilis)LAC4基因调控型启动子的控制之下，构建成h-FIX基因表达盒。将此表达盒插入克鲁维酵母的整合质粒，得到了一个高稳定的h-FIX基因整合表达载体pHKB202-FP。经摇瓶培养并以半乳糖诱导此表达载体转化的酵母菌株，ELISA检测显示酵母上清液中的人凝血因子IX表达量可达到约200ng/ml。此结果表明人凝血因子IX蛋白首次成功地在脆壁克鲁维酵母中得到了表达和分泌。这对于促进人凝血因子IX的基因工程产业化具有实际意义。

Expression and Secretion of Human Anti-haemophilic Factor IX in Yeast Kluyveromyces fragilis

A human anti-haemophilic factor IX gene (h-FIX) was fussed to a secretion signal peptide sequence from the yeast Kluyveromyces lactis and controlled under the regulatory promoter of LAC4 gene from K. fragilis to form an h-FIX gene expression cassette. By inserting the cassette into a yeast integrating plasmid, a high-stable h-FIX gene expression vector, pHKB202-FP, was constructed. After induction with galactose, the h-FIX was first successfully expressed and secreted in yeast K. fragils transformed by the pHKB202-FP. The expression level was up to 200 rig/ml in the culture supernatant determined by ELISA method. The result was significant to promotion of the h-FIX production in biotech industry scale.