WHY NO ONE REPLIES, is this a stupid question ? - (Jun/06/2014 )

Dear All, 

I am amplifying HA gene of influenza. I Have some late signals in the uninfected tissue ( no virus ) which differ from the know infected by 5-7 CT cycles but I have not any signals in the NTC ( meaning no primer dimer) nor in the -RT. I changed all stocks several time and the place of work and I am sure no contamination. 

the other strange thing is that in the positive sample and in these uninfected ones, I got signals in all concentrations of the primer ( 10, 7.5, 5, 2.5 picomole) I expected that the low concentration will not give any signals ? 

Any explanations ?

Thanks 

Hi Mohammed, 

I think the doubts you were having were pretty much clarified, in your previous discussion here

Probably what you are getting is noise coming out of some non specific binding of your primers in your Negative samples. Like others mentioned in your previous discussion, have you tried running that PCR product on a gel. What is its size? 

Note also that this is a volunteer forum - we don't just sit on here all day waiting for your questions, and we may not have answers to questions that you ask... getting angry is no way to get people to help you!

Dear Global moderator,

I think this will not take so much of time from you or from anyone else to say we answered this or we do not have extra solutions. This is more better and even nicely accepted than no answer.

Thanks

No - it doesn't take much time, but there are a lot of people on here, and if I don't know it, there may well be someone else who does, so I don't comment.  If everyone who reads any post adds a comment saying that they don't know -  it doesn't help you and just clogs up Bioforum's server.  Lack of replies doesn't indicate that no-one is responding, - look at how many people read your topic - (65 for this one when I posted) that's an indication that no-one knows.

Anyway so thanks for your effort.