中文摘要：

构建经基因工程改造的Ara h 2表达载体,表达并纯化该蛋白,鉴定其过敏原性。将花生主要过敏原Arah 2基因序列进行颠换,并将颠换后的序列进行合成,再将合成后的基因克隆到原核表达载体pET-32a（＋）上,然后转入Origami宿主表达菌中;利用Isopropylβ-D-1-Thiogalactopyranoside（IPTG）诱导表达;通过Ni2＋亲和层析纯化目的蛋白;Western blotting和ELISA鉴定该重组蛋白的过敏原性。测序结果表明合成后的序列成功整合到原核表达载体pET-32a（＋）上。重组的目的蛋白纯化后经SDS-PAGE鉴定,蛋白大小与理论值相符。Western blotting和ELISA结果均表明经基因工程改造的Ara h 2（F-Ara h 2）蛋白与重组的Ara h 2（R-Ara h 2）蛋白相比,结合花生过敏患者混合血清中IgE显著降低。成功构建经基因工程改造的Ara h 2表达载体,初步的体外实验表明该基因表达的重组蛋白具有低致敏原性。

英文摘要：

The genetically engineered major allergen Ara h 2 was expressed and purified and the hypoallergenic of purified recombinant F-Ara h 2 protein was characterized.The Ara h 2 was reassembled and inserted into the expression vector pET-32a（＋）.The vector was transformed into Origami and the protein expression was induced by IPTG.Ni2＋ Chelating affinity chromatography was used to purify the recombinant F-Ara h 2 protein.The hypoallergenic of F-Ara h 2 was examined by Western blotting and ELISA.The ORF which contained 471 bp and encoded 157 amino acids was authenticated to be F-Ara h 2.The recombinant F-Ara h 2 protein induced by IPTG was consistent with the actual value.The affinity between recombinant F-Ara h 2 protein and IgE antibodies from pooled peanut-allergic patients serum was significantly decreased compared with R-Ara h 2 was identified by Western blotting and ELISA.Recombinant F-Ara h 2 protein was hypoallergenic.