real time PCR / qPCR

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real time PCR / qPCR

Post by charlotte » Thu Apr 27, 2006 7:12 pm

I am new to the area of real time PCR and am hoping someone can point me in the right direction to start. I work with plants if that helps. Someone mentioned that I need to pick a houskeeping gene....What does this mean? I have some knowledge of the theory for real time PCR so I don't need an entire background but I have no idea where to start with my experiments since I've never done it.
Any help would be great!

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Post by LilKim » Fri Apr 28, 2006 3:11 am

qPCR has several applications.... so i'm not sure what would apply to your plant-work.

A housekeeping gene is a gene that you normalize your experiement to. This gene should have a known transcription level that will not increase or decrease in comparison to whatever your "experiemental" cDNA is. For example, if you were looking at the expression of a gene involved in a learning process in mice (gene X)... you'd challenge your mice to learn a maze.. thus, you'd expect gene X's transcription levels to increase ... however, you'd pick a "house keeping gene to normalize your sample". Commonly used genes are GAPDH or Actin... because these genes are always transcribed at a constant rate. (the transcription rate will not chage because you're making your mouse run through a maze)

However, i don't know much about i don't know of any good housekeeping genes. Maybe you should pubmed-it ... to see what other plant-people use?

good luck!

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