A copper-controlled RNA interference system for reversible silencing of target genes in Trichoderma reesei.
A copper-controlled RNA interference system for reversible silencing of target genes in Trichoderma reesei.
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用于里氏木霉靶基因可逆沉默的铜控制 RNA 干扰系统
DOI：
10.1186/s13068-018-1038-7
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发表时间：
2018
影响因子：
6.3
通讯作者：
Liu W
中科院分区：
文献类型：
--
作者：
Wang L;Zheng F;Zhang W;Zhong Y;Chen G;Meng X;Liu W
背景：里氏木霉是工业上主要的木质纤维素酶产生菌。然而，纤维素酶的合成以及其他生理过程的机制还没有得到充分的理解，部分原因是其遗传操作的复杂过程。RNA干扰（RNAi）是真核生物（包括丝状真菌）基因研究和生物技术的有力工具。以往报道的里氏木霉RNAi系统要么是不可控的，要么只适用于特定的营养状态。结果：本研究将铜响应tcu1启动子整合到RNAi介导的沉默系统中，构建了一个可调控的里氏木霉RNAi沉默系统。作为概念验证，当表达相应的RNAi片段时，原养型pyr4基因、由Avicel诱导的高表达的cel7a和xyr1基因和afab1基因（其敲除已被证明是难处理的）在铜不存在的情况下被成功敲除。重要的是，通过在培养基中添加铜，RNAi菌株的表型显示出容易逆转以模拟互补，从而排除由hpRNA盒的随机整合引起的任何不想要的效应。结论：铜控制的RNA干扰系统是一种有效的逆转录沉默靶基因的工具，这种有条件地敲低或关闭基因的策略将有助于我们对里氏木霉的了解。reeseigenes功能，特别是那些由于各种原因难以被敲除的功能。该方法简单、成本低，在合成生物学和基因工程中具有广阔的应用前景。reesei。
Background:Trichoderma reeseiis a primary lignocellulosic enzyme producer in industry. However, the mechanisms underlying cellulase synthesis as well as other physiological processes are insufficiently understood partly due to the sophisticated process for its genetic manipulation. Target gene knockdown by RNA interference (RNAi) is a powerful tool for genetic research and biotechnology in eukaryotes including filamentous fungi. Previously reported RNAi system inT. reeseiwas either uncontrollable or only applicable in certain nutrition state.Results:In the present study, we incorporated the copper-responsivetcu1promoter into an RNAi-mediated silencing system to develop a controllable RNAi-mediated silencing system inT. reesei. As the proof-of-concept, a prototrophicpyr4gene, highly expressedcel7aandxyr1genes induced by Avicel and afab1gene, whose knockout has proved to be intractable, were successfully knocked down in the absence of copper when the respective RNAi fragment was expressed. Importantly, the phenotype of RNAi strains was shown to be reversed easily to mimic the complementation for excluding any unwanted effects resulted from the random integration of the hpRNA cassette by adding copper in the media. Thus, this controllable RNAi-mediated silencing system can be turned on and turned off only depending on the absence and presence of copper ions in the media, respectively, and not on the nutritional states.Conclusions:The copper-controlled RNA interference system represents an effective tool for reversible silencing of target genes inT. reesei.This reported strategy to conditionally knock down or turn off genes will contribute to our understanding ofT. reeseigene functions, especially those that are difficult to be knocked out due to various reasons. In addition, this simple and cost-effective method holds great potential for the application in synthetic biology and genetic engineering ofT. reesei.
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影响因子：
6.3
作者：
Kubicek CP;Mikus M;Schuster A;Schmoll M;Seiboth B
通讯作者：
Seiboth B
影响因子：
16.6
作者：
Dong, Xian-ping;Shen, Dongbiao;Wang, Xiang;Dawson, Taylor;Li, Xinran;Zhang, Qi;Cheng, Xiping;Zhang, Yanling;Weisman, Lois S.;Delling, Markus;Xu, Haoxing
通讯作者：
Xu, Haoxing
影响因子：
6.4
作者：
Bischof RH;Horejs J;Metz B;Gamauf C;Kubicek CP;Seiboth B
通讯作者：
Seiboth B
铜响应启动子的表征及其介导的木聚糖酶调节剂 1 的过表达导致里氏木霉中不依赖于诱导的纤维素酶产生
影响因子：
6.3
作者：
Lv, Xinxing;Zheng, Fanglin;Liu, Weifeng
通讯作者：
Liu, Weifeng
DOI：
10.1083/jcb.143.1.65
发表时间：
1998-10-05
期刊：
The Journal of cell biology
影响因子：
--
作者：
Gary JD;Wurmser AE;Bonangelino CJ;Weisman LS;Emr SD
通讯作者：
Emr SD