I have qRT-PCR data of 4 transgenic line and 1 non-transgenic line. Non-trangenic line has GOI Ct value zero and HKG has Ct value because non-transgenic has no transgene/GOI, that's why no Ct value.
Please guide me how to analyze the data by delta delta Ct method.
Thanks,
Best wishes.
You are attempting to derive a 'fold change' value for a change of 'infinite increase' (zero >> literally anything).
Simply: don't do this.
Ask yourself what you actually want to show, and what you want to measure. If you are trying to show differences between the transgenic cell lines, then this can easily be done via dCt (while your non-transgenic cell line would simply be "zero", or more accurately: 'not detected').
Remember: unless you're obtaining absolute transcript numbers, all your expression values are relative. The actual numbers could be anything, it is simply the relationship between them that matters.
You are attempting to derive a 'fold change' value for a change of 'infinite increase' (zero >> literally anything).
Simply: don't do this.
Ask yourself what you actually want to show, and what you want to measure. If you are trying to show differences between the transgenic cell lines, then this can easily be done via dCt (while your non-transgenic cell line would simply be "zero", or more accurately: 'not detected').
Remember: unless you're obtaining absolute transcript numbers, all your expression values are relative. The actual numbers could be anything, it is simply the relationship between them that matters.
Dear Naveed
Actually, you can only use delta delta Ct method to compare between the lines that showed GOI expression and to calculate how many folds of difference is present between these lines.
As John mentioned you can use the absolute number of copies to compare between the lines with GOI expression and the non-transgenic line.
good luck
Dear Johan Hildyard and Michael Alaboud, Many Thanks for your valuable suggestion and comments.
Best Wishes.
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