人α-防御素-1基因与J链基因的融合及其表达载体的构建

Reconstruction of J Chain Gene with HNP-1 Gene and Construction of Its Expression Vector

目的本研究旨在将α-防御素-1(HNP-1)基因与J链基因进行融合使其成为融合基因J-HNP-1。方法从pCH-J质粒中扩增出J链;从pBabeNeo-HNP-1质粒中扩增出HNP-1;然后以它们的自身产物为模板,进行重组PCR,获取J-HNP-1 DNA片段,并插入编码双重报告基因Myc和6个多聚组氨酸(His)的哺乳动物细胞表达载体pcDNA3.1(-)/Myc-HisC中,构建出C端带Myc和6×His的rpcDNA3.1(-)/Myc-HisC/J-HNP-1。结果经过重组得到的融合基因J-HNP-1的目的片段为786 bp,与预测相符。结论杀菌分子J-HNP-1的重组和哺乳动物细胞表达载体的构建,为进一步进行抗菌肽的研究及制备奠定了基础。

OBJECTIVE To recombine J chain gene with HNP-1 into a new germicidal molecule J-HNP-1, which can connect with pIgR by J chain, so that by using plgR as a ＂bridge＂ the J-HNP-1 germicidal peptide can be transported into the epithelial cell of mucous membrane to kill the intraeellular microorganisms, then the recombinant is inserted into the mammalian expression system. METHODS The J chain and HNP-1 eDNA were amplified from the plasmids respectively by PCR, then the two eDNA fragments were reeombined into J-HNP-1 by recombinant PCR. The J-HNP 1 eDNA fragment was inserted into the mammalian expression vector pcDNA3. 1 （-）/MycHisC. RESULTS The J-HNP-1 recombinant was obtained by connection of J chain and HNP-1 eDNA by PCR. The recombinant J-HNP-1 eDNA was 786bp. CONCLUSIONS The recombinant J-HNP-1 eDNA and the construction of expression vector are the basis for the new bactericidal peptide production.