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Problems with RNA isolation

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#1 mts



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Posted 18 February 2010 - 08:31 AM

Hi everyone,

I isolated total RNA from a bacterial sample, using a commercial kit, in order to perform RT-PCR. I analysed the RNA extracted in an agarose gel and had 2 bands, as expected and apparently no cromossomal DNA. Before producing cDNA I did a control PCR wich revealed that the RNA I extracted was contaminated with DNA. So I did another treatment with DNase as described in that kit and checked the RNA on a gel again. Surprisingly one of the bands is missing, i.e. I can only observe the highest band. This band shows a good intensity, apparently there was no degradation of this form of RNA during the second extraction; simply the other band completely disappeared. I repeated the second extraction and the same happened. Iím not experienced in working with RNA and I was hoping to get some help solving this problem. Thanks!!!!

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