Forgive me if I missed an identical question here, but I couldn't find it.
I need to establish a TaqMan analysis for quantification of bacteria from various samples without culturing the bacteria on the media.
I've got everything except the internal amplification control (IAC) to check the false negative reactions.
As I understand there are several ways of developing an IAC, mainly the artificial construct cloned in a plasmid, DNA of another organism added to a reaction (together with specific primers and a probe) or a DNA fragment constructed in such a way, that the same primers are utilized for amplification as of a target gene.
My questions are as follows: 1. Which is the best and the quickest way to develop an IAC from your experience? 2. Are there any possibilities of ordering an already constructed, ready-made IAC for applications? If yes, which company offers them? [I have limited time] 3. Is it possible to use IAC in quantification reaction?
The co-amplification of another gene will always influence the yield of my target DNA, and in quantification real-time PCR I may not know the concentration of my target DNA so I won'y know the influence of IAC amplification on my target gene!
What's the solution?