摘要
用重组ＤＮＡ 技术研究了编码天麻抗真菌蛋白（ＧＡＦＰ）的基因，从天麻（Ｇａｓｔｒｏｄｉａ ｅｌａｔａＢｌ．）块茎中提取Ｐｏｌｙ（Ａ） ｍ ＲＮＡ 后合成ｃＤＮＡ，构建成表达型ｃＤＮＡ 文库，用纯化的蛋白质探针通过免疫筛选找出对应的ｃＤＮＡ 克隆。在进一步证明所选用的ｃＤＮＡ 克隆含有重组的λ－ｐｈａｇｅ ＤＮＡ 后，提取和纯化含有插入片段重组子的ＤＮＡ，用Ｅｃｏ ＲＩ酶切分析可见插入片段。
Antifungal protein is the main inhibitor of fungal infection in the secondary corm of Gastrodia elata Bl. was isolated and purified antifungal protein(GAFP) from the plant. Its molecular weight was about 14 kD. Polyclonal antibody against GAFP was produced. In vitro test, this antifungal protein inhibited the growth of some fungi in some crop including Gibberella zeae. cDNA was synthesized from poly(A) mRNA purified from G.elata. The cDNA was ligated into phage vector λgt11 DNA and packaged in vitro and the phages were propagated on E.coli Y1090 and a λgt11 expression library was constructed. A cDNA clone encoding for antifungal protein was screened out by immunoscreening of the library using the protein as a probe. The λDNA containing insert was digested by Eco RI after isolated and purified recombinants λDNA,the insert was obtained. The cDNA was 300 bp in length. The authors had isolated the cDNA clone encoding antifungal protein from G.elata.
关键词
天麻
抗真菌蛋白
CDNA
分子克隆
药用植物
Antifungal protein
Gene encoding antifungal protein
cDNA library
Protein probe
Immunoscreening
