宁夏苦豆子中产苦参碱内生真菌的分离与鉴定-论文.pdf

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1、中国农业科学2013，46(13)：2643．2654ScientiaAgriculturaSinicadoi：10．38648．issn．0578-1752．2013．13．003宁夏苦豆子中产苦参碱内生真菌的分离与鉴定余永涛，何生虎，赵清梅(宁夏大学农学院，银川750021；。北方民族大学生物科学与工程学院，银川750021)摘要：【目的】从宁夏野生苦豆子中分离内生真菌，筛选能够产生苦参碱、氧化苦参碱、氧化槐果碱或槐定碱4种喹诺里西啶类生物碱的内生真菌。【方法】应用光学显微镜和扫描电子显微镜对分离菌株显微形态进行观察，根据形态对各菌株进行初步分类；应用薄层色谱法、气相色谱一质

2、谱联用法和酸性染料比色法检测各分离菌株的生物碱成分。对能够产生生物碱的真菌ITS和SSUrDNA序列进行扩增和测序，根据序列信息进行同源性比较和系统发育树构建；结合形态特征，对其进行种属分类。【结果】从苦豆子中共分离到27株真菌，其中23株分属于青霉属、曲霉属、链格孢属、镰刀菌属、Simplicillium属真菌，4株为不产孢菌；筛选出E1、E3和E5这3株能够产生苦参碱的内生真菌，均属于Simplicillium属真菌，其菌丝中苦参碱的含量分别为20．4、17．4和37．6·g～。【结论】根据真菌形态特征、同源性比较和遗传进化分析结果，确定该3株产苦参碱内生真菌均为Simpli

3、cilliumlanosoniveLll1．关键词：苦豆子；苦参碱；内生真菌；SimplicilliumlanosoniveumIsolationandIdentificationofMatrine—ProducingFungalEndophytesfrOmSophoraalopecuroidesinNingxiaYUYong—tao，HESheng．hu，ZHAOQing—mei(CollegeofAgronomy,NingxiaUniversity,Yinchuan750021；CollegeofBiologicalScienceandEngineering,BeifangU

4、niversityofNationalities，~nchuan750021)Abstract：【0bjective】Anexperimentwasca~iedouttoisolateandconfirmwhetherthepresenceoffungalendophytethatcouldproducequinolizidinealkaloidsincludingmatrine，oxymatrine，oxysophocarpineorsophoridinefromwildSophoraalopeeuroidesLinNingxia．[Method]Theisolateswere

5、observedunderthelightmicroscopeandscanningelectronmicroscopy，respectively．Theclassificationofeachisolatewasperformedpreliminarilyaccordingtothemorphologicalcharacteristics．Thechemicalcompositionsofisolatedfungiweredetectedbythin-layerchromatography,gaschromatograph—massspectrometerandaciddyec

6、olorimetrytoscreenthequinolizidinealkaloids—producingfungalendophytes．TheITSandSSUrDNAsequencesofthefungiwereamplifiedandsequenced．Homologycomparisonwasconductedandphylogenetictreewasestablishedbasedonthesequencesofthefungi．Thealkaloids—producingfungalendophyteswereclassifiedaccordingtothefun

7、galmorphologyandphylogeneticanalysis．[Result]Twenty．sevenfungiwereisolatedfromSalopecuroides．Twenty—threeofthe27isolateswereclassifiedasPenicillium，Aspergillus，AlternariaandFusariumbasedonmorphologyofconidia．Theotherfourisolatescouldnotproduc