How can I isolate RNA from cell line suspended in RNAlater (I mean, how can I remove the RNAlater)? I use TRIzol for RNA isolation, does RNA later interfere with downstream processing (i.e. real time PCR)?
you can add chloroform and precipitate with isopropanol to get the RNA and use a PCR clean up kit before proceeding with RT-PCR.
http://www.invitrogen.com/etc/medialib/en/filelibrary/Nucleic-Acid-Purification-Analysis/PDFs.Par.18819.File.dat/bp_7020.pdf
page 3!
You can remove RNAlater by centrifugation, followed by a wash with filtered and autoclaved PBS. The trizol might be added on the pellet. Alternatively, if your cells are attached, it is better and easier to add the RNAlater on the plate, after removing the medium. When ypu decide to perform the extraction you may just do a rapid wash with PBS and then add the lysis solution.
@ Daniela lancu same steps I do but when I remove RNAlater using centrifugation I observe some sticky and waxy layer on tube...even after Chloroform wash some oily layers comes from supernatent previously I thought it is due to phenol contamination which may be comes from TRIzol. but I think this is due to RNA later.. Can any one suggest me what to do.???
http://www.qiagen.com/knowledge-and-support/faq/?ID=cf2ab174-ffc9-4d0c-890b-c9640b17fd2a
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