bacteriophage genome - (Jul/26/2012 )
Hello everyone, I'm trying to isolate the genome from a vibriophage and I'm having some problems. First i used a protocol based on fenol-chloroform extraction and ethanol precipitation, this protocol worked very well before with other phages that has dsDNA genome. This time with this protocol I just have a small band (less than 100 pb) of something that seems to be RNA. So I used a protocol of RNA extraction with Trizol, and this time I had three small bands of RNA (between 50 and 100 pb) (we have tried DNA and RNA viral isolation kits and the results is always the same). I'm sure is RNA because is sensitive to RNAse and resistant to DNAse treatment. These RNA is too small to be the genome of the phage. So there is anyone who has an Idea of what could be? I was thinking that maybe the genome is strongly attached to the protein phase or something, I'm not sure how to follow with this. Any help or suggestion?
is it a lytic, dsDNA phage? ...if so you could use kits used for lambda phage DNA purification.
It seems it's a virulent phage, we already tried with DNA purification kits. We think maybe belongs to leviviridae family since we isolate small RNA fragments, the problem is we can't see just a small band how should be with this kind of virus. Instead that we see small fragments.