玉米大斑病菌聚酮体合成酶基因 StPKS 功能分析 |
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引用本文: | 张鑫,董金皋,曹志艳,刘士伟,郭丽媛. 玉米大斑病菌聚酮体合成酶基因 StPKS 功能分析[J]. 中国农业科学, 2011, 44(8): 1603-1609. DOI: 10.3864/j.issn.0578-1752.2011.08.008 |
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作者姓名: | 张鑫 董金皋 曹志艳 刘士伟 郭丽媛 |
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作者单位: | 1. 河北农业大学;2. 河北,保定,河北农业大学毒素实验室;3. ; |
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摘 要: | 【目的】利用RNA干扰技术初步探讨玉米大斑病菌聚酮体合成酶基因(StPKS)的功能。【方法】本试验将玉米大斑病菌StPKS基因片段连接到pSilent-1载体中,构建StPKS基因的RNA干扰载体pSilent-StPKS1-2。通过聚乙二醇介导的方法转入玉米大斑病菌野生型菌株01-23的原生质体,利用潮霉素筛选得到转化子,采用半定量RT-PCR方法分析转化子StPKS基因的表达情况,显微观察转化子与野生型菌丝形态的差异。【结果】本研究构建的StPKS基因RNA干扰载体对沉默该基因表达是有效的,经潮霉素抗性筛选,得到了30株阳性转化子,对其中5株黑色素积累下降、菌落颜色变浅的转化子进行了半定量PCR分析发现,5株转化子的StPKS基因表达量均有所下降;显微观察发现5株转化子的菌丝形态很不规则,发生了膨大、变形、分枝等现象。【结论】StPKS基因在DHN黑色素合成途径中起作用,其表达量下降会减少黑色素的产生,试验同时发现该基因与菌丝形态密切相关。
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关 键 词: | 玉米大斑病菌 聚酮体合成酶基因 DHN黑色素 RNA干扰 |
收稿时间: | 2010-10-08; |
Obtaining and analysis on silent transformants of Polyketide synthase gene(StPKS) in Setosphaeria turcica |
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Abstract: | 【Objective】RNA interference technology was used to investigate the function of a polyketide synthase gene (StPKS) in Setosphaeria turcica.【Method】The silent vector, pSilent-StPKS1-2, was constructed by inserting StPKS fragments into plasmid pSilent-1 then trandformed into S. turcica 01-23 protoplasts through PEG. Transformants were selected on potato dextrose agar (PDA) containing hygromycin and the expression levels of StPKS was analysised by Semiquantitative RT-PCR. The hypae morphology of transformants and wide type was observed by microscope.【Results】The RNAi vector was effective to reduce the gene expression. Thirty hygromycin-resistant transformants were isolated and five melanin-deficient transformants were analysised by Semiquantitative RT-PCR. The results showed that StPKS mRNA was weakly expressed in white-color phenotypes. The transformants hypae morphology was anomalistic, including intumescent, anamorphic and ramose.【Conclusion】StPKS gene involved in the DHN-melanin biosynthesis and related to the mycelial morphology. The DHN-melanin biosynthesis was readuced when StPKS gene weakly expression. |
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Keywords: | Setosphaeria turcica Polyketide synthase gene DHN-melanin RNAi |
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