New Strains for Tissue-Specific RNAi Studies in Caenorhabditis elegans.
New Strains for Tissue-Specific RNAi Studies in Caenorhabditis elegans.
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线虫组织特异性RNAi研究的新菌株
DOI：
10.1534/g3.120.401749
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发表时间：
2020-11-05
期刊：
影响因子：
--
通讯作者：
Watts JL
中科院分区：
文献类型：
--
作者：
Watts JS;Harrison HF;Omi S;Guenthers Q;Dalelio J;Pujol N;Watts JL
RNA干扰是解剖基因功能的有力工具。在秀丽隐杆线虫中，摄取双链RNA会导致靶基因强烈的系统性敲低。通过组织特异性RNAi技术可以进一步了解基因功能。目前可用的组织特异性秀丽隐杆线虫菌株依赖于在RNAi缺乏遗传背景的组织或细胞中拯救RNAi功能。我们试图利用目前可用的组织特异性RNAi菌株评估特定组织对多不饱和脂肪酸（PUFA）合成的贡献。我们发现rde-1 （ne219）是一种常用的RNAi抗性突变株，对针对PUFA合成基因的RNAi保持相当大的RNAi能力。通过测量合成PUFAs的去饱和酶的脂肪酸产物的变化，我们发现前面提到的菌株rde-1 （ne219）和报道的仅种系RNAi菌株rrf-1 （pk1417）不适合进行组织特异性RNAi实验。然而，敲除突变体rde-1 （ne300）对dsRNA诱导的RNAi具有很强的抗性，因此更适合构建强大的组织特异性RNAi菌株。使用新构建的rde-1（零）背景菌株，我们发现在肠道、表皮和种系组织中有相当大的去饱和酶活性，但在肌肉中没有。本研究报道的rnai特异性菌株将为秀丽隐杆线虫研究人员研究各种生物过程提供有用的工具。
RNA interference is a powerful tool for dissecting gene function. In Caenorhabditis elegans, ingestion of double stranded RNA causes strong, systemic knockdown of target genes. Further insight into gene function can be revealed by tissue-specific RNAi techniques. Currently available tissue-specific C. elegans strains rely on rescue of RNAi function in a desired tissue or cell in an otherwise RNAi deficient genetic background. We attempted to assess the contribution of specific tissues to polyunsaturated fatty acid (PUFA) synthesis using currently available tissue-specific RNAi strains. We discovered that rde-1 (ne219), a commonly used RNAi-resistant mutant strain, retains considerable RNAi capacity against RNAi directed at PUFA synthesis genes. By measuring changes in the fatty acid products of the desaturase enzymes that synthesize PUFAs, we found that the before mentioned strain, rde-1 (ne219) and the reported germline only RNAi strain, rrf-1 (pk1417) are not appropriate genetic backgrounds for tissue-specific RNAi experiments. However, the knockout mutant rde-1 (ne300) was strongly resistant to dsRNA induced RNAi, and thus is more appropriate for construction of a robust tissue-specific RNAi strains. Using newly constructed strains in the rde-1(null) background, we found considerable desaturase activity in intestinal, epidermal, and germline tissues, but not in muscle. The RNAi-specific strains reported in this study will be useful tools for C. elegans researchers studying a variety of biological processes.
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