小花棘豆(Oxytropis glabra DC.)11株内生真菌的分离培养及鉴定

Isolation,Culture and Identification the 11 Endophytic Fungus from Oxytropis glabra DC.

用小花棘豆(Oxytropis glabra DC.)单株植物叶和茎切段做外植体,体外分离并培养内生真菌,研究菌落和分生孢子的微生物形态特征,对培养出的内生真菌的5.8SrDNA/ITS区进行扩增及序列测定,序列输入GenBank并与已发表的小花棘豆内生真菌序列进行同源性比对,并利用DNASTAR软件包中MegAlign软件构建系统发育树,结合分子生物学及微生物学研究推测内生真菌所属类群.结果显示:11株小花棘豆内生真菌的菌落和分生孢子的微生物形态特征与埃里砖格孢属真菌Embellisiasp.L12特征一致;小花棘豆内生真菌间ITS1和5.8SrDNA区序列相同,与Embellisiasp.L12相比只有一个变异位点,位于5.8SrDNA区,在ITS2区另有两个变异位点.推测小花棘豆内生真菌应与埃里砖格孢属亲缘关系密切,对其物种种属分类地位还须深入研究.

Leaves and stems were cut from individual plants of Oxytropis glabra DC. , and used as materials to isolate and culture the endophytic fungus in vitro. The microbiological morphology of colony and the conidiophore were investigated. The 5.8SrDNA/ITS of the in vitro cultured endophytic fungi were sequenced, and then the sequence data of the endophytic fungi were submitted to GenBank and analysed by homologous blast. The phylogenetic tree was constructed by MegAlign of DNASTAR software package. The molecular biological and microbiological characters were investigated in order to predict the classification of the endophytic fungi. The results indicated that the morphologic characteristics of the colony and conidiophore of this 11 endophytic fungi were the same as that of Embellisia sp. L12. The sequences of ITS1 and 5. 8SrDNA of endophytic fungus from Ozytropis glabra DC. were totally the same. Compared to the sequence of Embellisia sp. L12, there was only one viriation site which was in the 5.8SrDNA region in each sequence. In addition, two variation sites were shown in the ITS2 region. It was presumed that the endophytic fungi had a close relationship to Embellisia, but the classification of the endophytic fungi need further research.