Nova Reader - Subject https://www.novareader.co Default RSS Feed en-us Newgen KnowledgeWorks <![CDATA[The CONJUDOR pipeline for multiplexed knockdown of gene pairs identifies RBBP-5 as a germ cell reprogramming barrier in <i>C. elegans</i>]]> https://www.novareader.co/book/isbn/10.1093/nar/gkaa1171 Multiple gene activities control complex biological processes such as cell fate specification during development and cellular reprogramming. Investigating the manifold gene functions in biological systems requires also simultaneous depletion of two or more gene activities. RNA interference-mediated knockdown (RNAi) is commonly used in Caenorhabditis elegans to assess essential genes, which otherwise lead to lethality or developmental arrest upon full knockout. RNAi application is straightforward by feeding worms with RNAi plasmid-containing bacteria. However, the general approach of mixing bacterial RNAi clones to deplete two genes simultaneously often yields poor results. To address this issue, we developed a bacterial conjugation-mediated double RNAi technique ‘CONJUDOR’. It allows combining RNAi bacteria for robust double RNAi with high-throughput. To demonstrate the power of CONJUDOR for large scale double RNAi screens we conjugated RNAi against the histone chaperone gene lin-53 with more than 700 other chromatin factor genes. Thereby, we identified the Set1/MLL methyltransferase complex member RBBP-5 as a novel germ cell reprogramming barrier. Our findings demonstrate that CONJUDOR increases efficiency and versatility of RNAi screens to examine interconnected biological processes in C. elegans with high-throughput.

CONJUDOR adapts bacterial conjugation to combine RNAi plasmids in bacteria for robust knock-down of multiple genes in the nematode C. elegans. CONJUDOR has been used for RNAi screening to deplete multiple gene pairs with high-throughput in order to identify barriers of germ cell reprogramming into GABAergic motor neurons. The Set1/MLL methyltransferase complex member RBBP-5 could be identified using CONJUDOR as a novel barrier for germ cell to neuron reprogramming.
Graphical Abstract
CONJUDOR adapts bacterial conjugation to combine RNAi plasmids in bacteria for robust knock-down of multiple genes in the nematode C. elegans. CONJUDOR has been used for RNAi screening to deplete multiple gene pairs with high-throughput in order to identify barriers of germ cell reprogramming into GABAergic motor neurons. The Set1/MLL methyltransferase complex member RBBP-5 could be identified using CONJUDOR as a novel barrier for germ cell to neuron reprogramming.

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