Cloning And Expression On Secretive Proteins Anx-1of P.infestans And Vector Construction Of Gene Pipme1

Potato （Solanum tuberosum L.） is the fourth largest food crops in both the worldand China. Late blight is a major disease in potato cultivation, which caused by theoomycete Phytophthora infesrans（Mont.）de Bary. The research from the angle of themolecules on pathogenic gene of Phytophthora infesrans is a research hotspot inrecent years.The laboratory previous studies have established a system for metabolitesextraction and purification of P.infestans. Sixteen protein sequences was abtained byLC-MS, which including two annexin family. One of the annexin family was selectedto do continue research in order to clear the role of the enzyme in the disease process.A green fluorescent protein eukaryotic expression vector for gene pipme1wasconstructed which toxicity was got and clear.Total RNA from the mycelium of strain LSX18was extracted using TRIZOLreagent, the primes was designed with the nucleotide sequence of XP₀02897238. Aannexin family gene named anx-1（KJ624634） was cloned by the primes. The fμlllength was987bp, which contained a987bp ORF and encoding a protein of328amino acids. The theoretical molecular weight of protein was36.211kDa and IP was5.54.The anx-1was prokaryotic expressed by Escherichia coli, the recombinantprotein was inoculated on the potato leaves in vitro, while the similar necrotic spots asthe symptoms of potato late blight was not appeared on the tested leaves. Therefore,the expression protein of anx-1was not one of the factors that caused leaded to tissuenecrosis in potato leaves. Meanwhile, a green fluorescent protein eukaryotic expression vector wassuccessfully constructed according to the known virose pipme1gene sequence. Theresearch will help to deeply study on the molecular mechanisms of the pathogen andhost interaction and provide the basis for potato late blight pathogenic mechanism.