The Cloning And Expression Analysis Of Gmhinl Gene Related To Phytophthora Sojae Resistance

Phytophthora root and stem rot of soybean caused by Phytophthora sojae Kaufmann&Gerdemann is a destructive disease endangering soybean production all over the world.Plants can be induced to hypersensitive reaction (HR) by Harpin protein, and HIN1(harpin-induced1) is one of the harpin induction of protein expression in resistance to pathogen attack. There has been no report on HINI gene in soybean till now. In previous report, a cDNA library enriched for mRNAs encoding ESTs that increased in abundance during infection with Phytophthora sojae was constructed by suppression subtractive hybridization (SSH) coupled with cDNA microarrays from leaf tissues of high resistant soybean’Suinong10’, and an EST homologous to AtNHL3(GenBank accession no.BT000871.1) showed differential abundance in response to P. sojae infection. The objective of this study was to clone Hinl gene by RT-PCR from resistant soybean’Suinong10’, and the function of this gene was preliminarily analyzed.The main results were as following:1. The full-length of Hinl gene (designated as GmHinl) was871bp containing a651bp open reading frame which encoded a216residue polypeptide with a calculated molecular weight of24.48kDa and isoelectric point (PI) of9.63. Hinl was a hydrophilic protein, and contained a conserved domain of LEA-2. Sequence comparison showed that the putative protein was highly homologous to that from tobacco NtHIN1(57%), tobacco NtHIN2(56%), potato StHIN1(55%).2. Real-time PCR was conducted to examine the expression pattern of GmHinl, and the results showed that GmHinl was constitutively and highly expressed in the leaves, followed by roots and stems. The mRNA transcript was increased with treatments of P. sojae, jasmonate (JA) salic acid (SA), and gibberellin (GA3), and decreased with treatment of abscisic acid (ABA) The expression of GmHinl was also induced with treatments of drought, mechanical damage and low temperature.3. GmHinl was found localized to the plasma membrane with confocal laser microscope.4. The plant expression vectors of GmHinl was constructed and transformed into tobaccos to overexpress the gene by way of Agrobacterium tumefaciens mediation. We gained19resistant plants.Compared with non-transgenic tobacco, transgenic tobacco has a higher resistance.