金线莲分泌IAA内生真菌的筛选与鉴定及其发酵条件优化

Screening and Identification of Endophytic Fungi Secreting IAA in Anoectochilus roxburghii and Optimization of Fermentation Conditions

从福建金线莲品种‘红霞’的根、茎、叶中分离内生真菌,分析优良菌株产吲哚乙酸(IAA)的特性.采用组织培养、平板划线法和Salkowski比色法,筛选能够产IAA的金线莲内生真菌,通过形态特征及内部转录间隔区(internal transcribed spacer,ITS)序列等方法进行菌种鉴定,并利用正交试验确定菌种产IAA最优条件.结果表明:从‘红霞’金线莲茎中筛选分离的内生真菌PJ3产IAA能力最强,产量高达106.7μg·mL^(-1);经鉴定,PJ3为角担菌属(Ceratobasidium)真菌.PJ3最佳发酵培养基配方和培养条件分别为:0.5 g·L^(-1)硫酸镁,1.5 g·L^(-1)磷酸氢二钾,1.5g·L^(-1)磷酸二氢钾,10.0 g葡萄糖,15.0 g酵母浸粉,培养时间为132 h,培养温度为28℃,初始pH值为5,摇床转速180 r·min^(-1),装液量(体积分数)为28%.

This study aims to analyze the characteristics of indole acetic acid(IAA) secretion in excellent strains of ‘Hongxia’ Anoectochilus roxburghii by isolating the endophytic fungi from the roots,stems and leaves,thus exploring the resources of excellent growth promoting strains.Endophytic fungi of ‘Hongxia’ Anoectochilus roxburghii that could secret IAA were screened out by culturing,streak plate method and Salkowski colorimetric method,and then identified by observing morphological characteristics and internal transcribed spacer(ITS) sequencing.Finally,the optimal conditions of IAA secretion were determined by the orthogonal test.The results show that the endophytic fungus PJ3 isolated from the stems of ‘Hongxia’ Anoectochilus roxburghii presented the strongest ability to secrete IAA,and the secretion amount was up to 106.78μg·mL^(-1).PJ3 was identified as the Ceratobasidiumfungus.The optimal fermentation medium formula and culture conditions of PJ3 were as follows:magnesium sulfate is 0.5g·L^(-1),dipotassium hydrogen phosphate is 1.5g·L^(-1),potassium dihydrogen phosphate is 1.5g·L^(-1),glucose is 10.0g,yeast extract powder is 15.0g,culture time is 132h,culture temperature is 28 ℃,initial pH value is 5,rotation speed is 180r·min^(-1),and the liquid volume(volume fraction)is 28%.