利用RACE技术从板栗短雄花序突变体中扩增得到660bp的板栗脂质转运蛋白(lipid transfer protein,LTP)cDNA片段。该cDNA编码118个氨基酸,具有8个位置保守的半胱氨酸(C)残基及26个氨基酸的信号肽,它与棉花、草莓脂质转运蛋白氨基酸序列相似性为63%,基因序列已提交数据库(GenBank),其登录号分别为FJ490676(基因)和ACL01093(蛋白)。荧光定量分析表明板栗短雄花序突变体较正常雄花序的脂质转运蛋白表达量更高。将板栗脂质转运蛋白基因插入到硫氧还蛋白融合表达载体pET32a(+)中,构建了板栗的原核表达载体pET-LTP,在大肠杆菌菌株Rosetta-gamiTM2(DE3)中,IPTG诱导5h大量表达约为30ku的融合蛋白,通过Ni2+-chelating sepharose fast flow柱纯化的融合蛋白具有抑制板栗镰刀菌属病原真菌孢子萌发的功能。
rDNA internal transcribed spacer analysis(rDNA ITS)was used to study diversity of 12 representative mycorrhizal Rhizoctonia strains,which were isolated from Chinese orchids(Cymbidium)distributed in different sites and belonged to different ecologic types.The Blast results indicated that all these strains belonged to Epulorhiza or Tulasnella,which was fully consistent with identification as Epulorhiza with morphological method.Additionally,based on rDNA ITS sequence cluster analysis,dendrogram of Cymbidium mycorrhizal strains showed that the distributed environments and orchids species were two crucial factors that affected the specificity-relation between Chinese orchids(Cymbidium)symbiotic mycorrhizae and hosts.
