对RNAi是否做rescue experiments补充
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试验整体方案
1.Scrambled (Negative) siRNA Control
2.Positive siRNA Control
3.Multiple siRNAs to a Single Target
4.Monitor Both Target mRNA and Protein Levels
5.Use the Lowest Possible siRNA Concentration
6.Validation Via Array Analysis and Rescue Experiments
To better understand an siRNA's effect, global gene expression patterns can be analyzed (e.g. using microarray analysis). Differential expression patterns can be compared between non-transfected cells, cells transfected with a negative control siRNA, and cells transfected independently with 2 or more gene specific siRNAs. Yes, it is time consuming and can be expensive, but this ultimate control gives the best indication of specific versus non-specific siRNA effects.
To better understand an siRNA's effect, global gene expression patterns can be analyzed (e.g. using microarray analysis). Differential expression patterns can be compared between non-transfected cells, cells transfected with a negative control siRNA, and cells transfected independently with 2 or more gene specific siRNAs. Yes, it is time consuming and can be expensive, but this ultimate control gives the best indication of specific versus non-specific siRNA effects.
Rescue experiments in which the RNAi effect is reversed through expression of a target gene refractory to silencing by a particular siRNA can give final confirmation that an observed effect is specifically due to knocking down the target of interest. Like array analysis, this experiment is time consuming.
While considered the ultimate controls, editors of Nature Cell Biology and other journals do not require these experiments for publication in their journals.
1.Scrambled (Negative) siRNA Control
2.Positive siRNA Control
3.Multiple siRNAs to a Single Target
4.Monitor Both Target mRNA and Protein Levels
5.Use the Lowest Possible siRNA Concentration
6.Validation Via Array Analysis and Rescue Experiments
To better understand an siRNA's effect, global gene expression patterns can be analyzed (e.g. using microarray analysis). Differential expression patterns can be compared between non-transfected cells, cells transfected with a negative control siRNA, and cells transfected independently with 2 or more gene specific siRNAs. Yes, it is time consuming and can be expensive, but this ultimate control gives the best indication of specific versus non-specific siRNA effects.
To better understand an siRNA's effect, global gene expression patterns can be analyzed (e.g. using microarray analysis). Differential expression patterns can be compared between non-transfected cells, cells transfected with a negative control siRNA, and cells transfected independently with 2 or more gene specific siRNAs. Yes, it is time consuming and can be expensive, but this ultimate control gives the best indication of specific versus non-specific siRNA effects.
Rescue experiments in which the RNAi effect is reversed through expression of a target gene refractory to silencing by a particular siRNA can give final confirmation that an observed effect is specifically due to knocking down the target of interest. Like array analysis, this experiment is time consuming.
While considered the ultimate controls, editors of Nature Cell Biology and other journals do not require these experiments for publication in their journals.
