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Clone mapper: an online suite of tools for RNAi experiments in Caenorhabditis elegans.

Thakur N, Pujol N, Tichit L, Ewbank JJ - G3 (Bethesda) (2014)

Bottom Line: Proper interpretation of results from RNAi experiments requires a series of analytical steps, from the verification of the identity of bacterial clones, to the identification of the clones' potential targets.Despite the popularity of the technique, no user-friendly set of tools allowing these steps to be carried out accurately, automatically, and at a large scale, is currently available.We show that Clone Mapper overcomes the limitations of existing techniques and provide examples illustrating its potential for the identification of biologically relevant genes.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie de Marseille-Luminy, UM2 Aix-Marseille Université, Case 906, 13288 Marseille Cedex 9, France INSERM U1104, 13288 Marseille, France CNRS UMR7280, 13288 Marseille, France.

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Network analysis of novel RNAi targets. (A) Ceron et al. undertook an RNAi screen to identify genes that interact with the C. elegans retinoblastoma gene lin-35 (Ceron et al. 2007). The list of novel targets identified with Clone Mapper for the RNAi clones selected by Ceron et al. was used as input to GeneMania (black circles), together with lin-35/C32F10.2 (highlighted in yellow) as a seed gene. (B) Fievet et al. performed RNAi screens for C. elegans cell polarity mutants, to generate a polarity network (Fievet et al. 2013). A list of novel targets identified with Clone Mapper for the RNAi clones used by Fievet et al. was used as input to GeneMania (yellow circles), together with the genes corresponding to the 14 mutant strains used in the study (black circles). (C) Roy et al. performed a screen to find components of a regulatory network that promotes developmentally programmed cell-cycle quiescence (Roy et al. 2014). Novel targets identified with Clone Mapper for the RNAi clones used by Roy et al. (yellow), together with common targets (black) were used as input to GeneMania. The networks were trimmed to retain only direct neighbors; unconnected genes are not shown. Genes that are linked within GeneMania but do not appear on the list of RNAi clone targets are shown as gray circles; their size is proportional to the calculated probability score. Networks were displayed in Cytoscape; green edges represent experimentally-determined genetic interactions, pink edges represent experimentally-determined physical interactions for the corresponding proteins, orange and gray edges interactions predicted on the basis of co-expression or literature mining, respectively.
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fig6: Network analysis of novel RNAi targets. (A) Ceron et al. undertook an RNAi screen to identify genes that interact with the C. elegans retinoblastoma gene lin-35 (Ceron et al. 2007). The list of novel targets identified with Clone Mapper for the RNAi clones selected by Ceron et al. was used as input to GeneMania (black circles), together with lin-35/C32F10.2 (highlighted in yellow) as a seed gene. (B) Fievet et al. performed RNAi screens for C. elegans cell polarity mutants, to generate a polarity network (Fievet et al. 2013). A list of novel targets identified with Clone Mapper for the RNAi clones used by Fievet et al. was used as input to GeneMania (yellow circles), together with the genes corresponding to the 14 mutant strains used in the study (black circles). (C) Roy et al. performed a screen to find components of a regulatory network that promotes developmentally programmed cell-cycle quiescence (Roy et al. 2014). Novel targets identified with Clone Mapper for the RNAi clones used by Roy et al. (yellow), together with common targets (black) were used as input to GeneMania. The networks were trimmed to retain only direct neighbors; unconnected genes are not shown. Genes that are linked within GeneMania but do not appear on the list of RNAi clone targets are shown as gray circles; their size is proportional to the calculated probability score. Networks were displayed in Cytoscape; green edges represent experimentally-determined genetic interactions, pink edges represent experimentally-determined physical interactions for the corresponding proteins, orange and gray edges interactions predicted on the basis of co-expression or literature mining, respectively.

Mentions: To evaluate the potential impact of these differences in prediction, we compared the list of putative targets in four published data sets with those obtained with Clone Mapper. In the first screen, where just 29 clones were selected (Pukkila-Worley et al. 2014), Clone Mapper predicted the same targets as published; no novel targets with high scores were identified. In the second specific case (Ceron et al. 2007), 14 of 244 targets were not predicted by Clone Mapper since the insert sequences of the corresponding clones cannot be predicted. On the other hand, Clone Mapper identified 23 new targets with of score >1, 9 of which had a score >50 (Supporting Information, Table S1). Similar results were obtained for the two other studies (Fievet et al. 2013; Roy et al. 2014) (Table 1, Table S2, and Table S3). In all cases, the novel targets identified with Clone Mapper formed part of a closely linked network (Figure 6). The interconnectivity of the novel RNAi targets suggests that they may be functionally important for the biological process under study. Such a hypothesis requires direct experimental validation, but the results demonstrate the potential utility of Clone Mapper in gene discovery.


Clone mapper: an online suite of tools for RNAi experiments in Caenorhabditis elegans.

Thakur N, Pujol N, Tichit L, Ewbank JJ - G3 (Bethesda) (2014)

Network analysis of novel RNAi targets. (A) Ceron et al. undertook an RNAi screen to identify genes that interact with the C. elegans retinoblastoma gene lin-35 (Ceron et al. 2007). The list of novel targets identified with Clone Mapper for the RNAi clones selected by Ceron et al. was used as input to GeneMania (black circles), together with lin-35/C32F10.2 (highlighted in yellow) as a seed gene. (B) Fievet et al. performed RNAi screens for C. elegans cell polarity mutants, to generate a polarity network (Fievet et al. 2013). A list of novel targets identified with Clone Mapper for the RNAi clones used by Fievet et al. was used as input to GeneMania (yellow circles), together with the genes corresponding to the 14 mutant strains used in the study (black circles). (C) Roy et al. performed a screen to find components of a regulatory network that promotes developmentally programmed cell-cycle quiescence (Roy et al. 2014). Novel targets identified with Clone Mapper for the RNAi clones used by Roy et al. (yellow), together with common targets (black) were used as input to GeneMania. The networks were trimmed to retain only direct neighbors; unconnected genes are not shown. Genes that are linked within GeneMania but do not appear on the list of RNAi clone targets are shown as gray circles; their size is proportional to the calculated probability score. Networks were displayed in Cytoscape; green edges represent experimentally-determined genetic interactions, pink edges represent experimentally-determined physical interactions for the corresponding proteins, orange and gray edges interactions predicted on the basis of co-expression or literature mining, respectively.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig6: Network analysis of novel RNAi targets. (A) Ceron et al. undertook an RNAi screen to identify genes that interact with the C. elegans retinoblastoma gene lin-35 (Ceron et al. 2007). The list of novel targets identified with Clone Mapper for the RNAi clones selected by Ceron et al. was used as input to GeneMania (black circles), together with lin-35/C32F10.2 (highlighted in yellow) as a seed gene. (B) Fievet et al. performed RNAi screens for C. elegans cell polarity mutants, to generate a polarity network (Fievet et al. 2013). A list of novel targets identified with Clone Mapper for the RNAi clones used by Fievet et al. was used as input to GeneMania (yellow circles), together with the genes corresponding to the 14 mutant strains used in the study (black circles). (C) Roy et al. performed a screen to find components of a regulatory network that promotes developmentally programmed cell-cycle quiescence (Roy et al. 2014). Novel targets identified with Clone Mapper for the RNAi clones used by Roy et al. (yellow), together with common targets (black) were used as input to GeneMania. The networks were trimmed to retain only direct neighbors; unconnected genes are not shown. Genes that are linked within GeneMania but do not appear on the list of RNAi clone targets are shown as gray circles; their size is proportional to the calculated probability score. Networks were displayed in Cytoscape; green edges represent experimentally-determined genetic interactions, pink edges represent experimentally-determined physical interactions for the corresponding proteins, orange and gray edges interactions predicted on the basis of co-expression or literature mining, respectively.
Mentions: To evaluate the potential impact of these differences in prediction, we compared the list of putative targets in four published data sets with those obtained with Clone Mapper. In the first screen, where just 29 clones were selected (Pukkila-Worley et al. 2014), Clone Mapper predicted the same targets as published; no novel targets with high scores were identified. In the second specific case (Ceron et al. 2007), 14 of 244 targets were not predicted by Clone Mapper since the insert sequences of the corresponding clones cannot be predicted. On the other hand, Clone Mapper identified 23 new targets with of score >1, 9 of which had a score >50 (Supporting Information, Table S1). Similar results were obtained for the two other studies (Fievet et al. 2013; Roy et al. 2014) (Table 1, Table S2, and Table S3). In all cases, the novel targets identified with Clone Mapper formed part of a closely linked network (Figure 6). The interconnectivity of the novel RNAi targets suggests that they may be functionally important for the biological process under study. Such a hypothesis requires direct experimental validation, but the results demonstrate the potential utility of Clone Mapper in gene discovery.

Bottom Line: Proper interpretation of results from RNAi experiments requires a series of analytical steps, from the verification of the identity of bacterial clones, to the identification of the clones' potential targets.Despite the popularity of the technique, no user-friendly set of tools allowing these steps to be carried out accurately, automatically, and at a large scale, is currently available.We show that Clone Mapper overcomes the limitations of existing techniques and provide examples illustrating its potential for the identification of biologically relevant genes.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie de Marseille-Luminy, UM2 Aix-Marseille Université, Case 906, 13288 Marseille Cedex 9, France INSERM U1104, 13288 Marseille, France CNRS UMR7280, 13288 Marseille, France.

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Related in: MedlinePlus