i have been doing qPCR analysis for some Pathogenicity related genes in Fungus but when i start calculating the fold changes using given protocol. the expression of control become 1 always. can anybody explain this phenomina to me ???
You say you calculate a fold change. This is the ratio between two groups, usually of an experimental group to a control group. If you talk about the fold change of the "control", do you mean the ratio of the control group to the control group? That must be 1, by definition. It is not particularily smart to to calculate a fold change between a group and itself.
Because through this analysis you explain how much time is more or less express your gene in the samples than the control. For example, the sample had 5 fold change of the gene A, means that this gene in your sample is 5 time more express than the control. And for example if is -3 means that the sample had 3 time less express on the sample than the control.
You say you calculate a fold change. This is the ratio between two groups, usually of an experimental group to a control group. If you talk about the fold change of the "control", do you mean the ratio of the control group to the control group? That must be 1, by definition. It is not particularily smart to to calculate a fold change between a group and itself.
The fold change of test group to control group?. If you're on this line, then no increase in the expression level of your test group relative to the controls. As you sounded, "the expression of control", not really clear because it has been expected to talk of your test group having already known that of the control, Again, if the expression you referred to was right and it is related to pathogenicity then, there is not higher risk of pathogenicity having ratified that the control gene was that of non-pathogenic organism, or mutant (as the case may be).
Giorgia Pertile do know any method by which we can calculate the expression ?
Ayaz Farzand https://www.youtube.com/watch?v=Kkle8T7aXjk this is the methodology of delta Ct method
By "....the expression of the control in qRT-PCR analysis always equal to 1", I assume that you intended to mean the expression of the reference gene (for instance wild-type). Normally, when you use the delta-delta CT method or any other method to analyze the qRT-PCR data, you will end up using the expression profile of control gene (e.g WT) to normalize the expression of other target genes, hence it becomes the reference expression level. The value 1 is a result of dividing the expression level value of the reference gene by itself in the process of normalizing the expression of other genes. For instance, if the expression level of your reference gene (control) is 8.3231218 and the expression levels of other two target genes (say A & B) are 12.0142812 and 6.1111102 , then upon normalization, the you will end up with the following values as the expression profiles of the control, gene A and gene B; 1.0000, 1.44348256, and 0.73423294 respectively.
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