(no subject)

[Dr. Hiranya S. Roychowdhury]

It is hard to infer the cause of your PCR failure from the info you have 
provided. The denaturation and annealing temps do not seem out of the ordinary, 
and 120 pMole of each primer (I asume that's what you meant) is plenty too.  

My questions:  How old are your primers? How did you reconstitute your primers 
and how did you store them?  I am asuming that you got good yield with this 
same batch of primers; in which case, my first suspect would be the integrity 
of the primers.  


Quoting sheeja At spices.res.in:

> dear all
> When i am trying to amplify the 16 s rRNA gene (1.5 kb), I am sometimes
> getting it while i am sometimes not. the bands are very faint on loading 5
> microlitre of pcr product. i used to get nice thick bands earier, but not
> now, i use 12o pm primer each, and all other std conditions for PCR, the
> denaturation temp is 92 for 2 min and annealing is 55 for 1 min. is it
> okay
> please some one suggest me a good way out
> thanks in advance
> tes
> 
> 
> 
> 
> 
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-- 
Hiranya S. Roychowdhury, Ph.D.
Asst. Professor,
Dept. of Health and Public Services, Rm# 191T,
Dona Ana Community College of New Mexico State University
MSC 3DA,
Las Cruces, NM 88003