Hello,
Can someone help me with any protocol to extract RNA from samples stored in ENAT medium (Copan) using TRIZOL?
Can I use directly the resuspended sample into Trizol using sample:Trizol proportion described by the manufacturer? (250 ul sample + 750ul TRIZOL)
Thanks
Hi Jorge,
I have read that, eNat is guanidine thyocianate based medium. Since the Trizol is based on this compound, they are compatible. So you can use your resuspended sample followed by Trizol addition.
The Trizol or any similar acid-guanidin-isothyocianate-phenol based lysis buffer effectively inhibit RNases in your samples. Then use the classical phenol-chloroform-isopropanol purifing method.
Alternatively you can use the Trizol lysate directly in Direct-zol RNA spin column kit https://www.zymoresearch.com/media/amasty/amfile/attach/_R2060_R2061_R2062_R2063_Direct-zol_RNA_MicroPrep_ver._1.0.0.pdf). I have very good experiences with this kit in human chordoma tissue RNA isolation in the terms of RNA quality and quantity. I have used the Trizole lysate after chloroform phase-separation step in the above kit. To decrease the protein contamination in the further purification steps
GITC/acidic phenol lysis combined with silica-based column methods
(Pros: after phase separation step no need to use chemical hood, can be high throughput Cons: more expensive than GITC-based methods
GITC/acidic phenol methods
(Pros: the cheapest, intact total RNA (including rRNAs, miRNAs) can isolated with high yield, Cons: toxic organic solvents (phenol, chlorophorm) (need a chemical hood), very problematic to use in high throughput).
Bests,
Tamas
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