血管生长抑制因子基因的克隆及其在昆虫细胞中的表达

目的；获得具有高活性的Ａｎｇｉｏｓｔａｔｉｎ表达，为进一步研究及应用奠定物质基础。方法：用ＰＣＲ法人纤维蛋白酶原ｐｌａｓｍｉｎｏｇｅｎ基因中，获得Ａｎｇｉｏｓｔａｔｉｎ（ｋｒｉｎｇｌｅｌ－４）基因，测序验证后，将正确的Ａｎｇｉｏｓｔａｔｉｎ（Ｋ１－４）序列，插入Ｂａｃ－ｔｏ－Ｂａｃ杆状病毒表达系统，转染昆虫细胞ｓｆ９，表达Ａｎｇｉｏｓｔａｔｉｎ。结果：获得了正确序列Ａｎｇｉｏｓｔａｔｉｎ ｃＤＮＡ

Clone and expression of angiostatin in insect cells

AIM: To have angiostatin expressed in sf9 insect cells by BactoBac baculovirus expression vector system. METHODS: Angiostatin( K14) cDNA was acquired by PCR from human plasminogen cDNA. First, angiostatin cDNA was inserted into pFast Bac vector, then the recombinant pFast Bacangio(K14) was transfected into competent DH10 Bac cells. Recombinant angio(K14) Bacmid DNA was constructed and sf9 cells were cotransfected with recombinant angio( K14)Bacmid DNA using cellfectin. RESULTS: After 3 round amplification of recombinant baculovirus, high expression level of angiostatin(K14) in sf9 cells infected was detected by immunoblotting with plasminogen(K14) antibody. The inhibitive activity was assayed using BALB/C 3T3 embryo cells. CONCLUSION: ZActive angiostatin was expressed in insect cells.