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When overexpressing several genes using different vectors, what negative control

over expression negative control empty vector experimental design gene expression

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#1 Lizzyf23

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Posted 30 October 2017 - 06:16 AM

Hi,

 

I'm planning on overexpressing six genes in two different cell lines, but the vectors all have different backbones, and they have 2 different resistance markers.

 

Can I use one empty vector negative control for all the vectors, or do I need a different empty vector control for each backbone? Alternatively, can I use an empty vector control for the 2 antibiotic resistance markers even though the backbone is different?

 

Thanks!



#2 bob1

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Posted 30 October 2017 - 06:32 AM

The answer depends on what you plan to do with the cell lines - i.e. what you want to measure during the experiment. Stringent controls would require you to use empty (or backbones containing scrambled target sequence) backbones in the same combinations and amounts (and ratios) as you do for the over-expression vectors. The same applies for the antibiotic resistance markers. One solution would be to subclone the genes all into the same backbone if possible, or if you are selecting for the presence of two different genes, then put some on one antibiotic and some on the other.



#3 Lizzyf23

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Posted 31 October 2017 - 10:42 AM

thanks for your answer - I suspected that but was hoping for a shortcut!







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