4 replies to this topic

[ACN]

I am isolating Sendai virus  mRNAs, from LLC-MK2 cells, and then quantifying using RT-PCR. I have used Roche's mRNA isolation kit (using magnetic beads); however I do not find the kit to be useful, as I run out the beads fairly quickly (and i'm not very trustful of reusing previously used beads). I was wondering if there are any suggestions for: Best mRNA isolation kits? (getting a lot of use out of it)

Thanks!

[jerryshelly1]

I have had excellent results with Ribozol.  Cheap and efficient.

[ACN]

Thanks for the info! Will this work for specifically isolating mRNAs and not total RNA ?

[jerryshelly1]

No total RNA.  Why do you need to isolate only mRNA?  If you are doing RT-PCR, it will not matter what RNA is isolated.  It will only rely on your GOI from your RT cDNA.  You must know the viral sequence of your gene to perform RT-PCR.  Are the viral primers you are using to similar to your LLC cells?  Maybe I am missing something.   If you absolutely need only mRNA, try the ambion mRNA pure kit from invitrogen.

[ACN]

Yeah, you're making a lot of sense! I all ready have the specific viral primers, used for the RT-PCR, but I am not sure if they are too close to the host cells. I actually never questioned why I can't just use total RNA, I'll bring it up to my P.I. tomorrow for sure.
Thanks for the help!