TRIzol Reagent
RNA isolation / purification Tissue - Human Intestine
Protocol tips
| Protocol tips |
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| - For low 260/230 readings the best approach is to try washing sample (precipitation) with ethanol to desalt it. |
Publication protocol
Small intestinal biopsies were stored in RNA stabilization solution RNAlater® (Ambion, Life Technologies, Carlsbad, CA) post assay and for total RNA purification lysed in 400 μl of TRIzol (Invitrogen, Life Technologies, Carlsbad, CA). Chloroform (80 μl) was added and whirl mixed for 15 sec. Homogenates were incubated 2–3 min at RT and centrifuged for 15 min at 10,000g (4°C). The water phase containing the RNA was further purified using RNeasy MinElute Cleanup Kit (Qiagen, Venlo, Netherlands). RNA concentration measured on a nano-drop 2000 (Thermo Scientific) and the integrity was confirmed on an Agilent 2100 Bioanalyzer using total RNA nano chips (Agilent Technologies, Santa Clara, CA, USA). RNA analysis of explant biopsies from 8 CD patients were performed, grading the quality post assay to prove cell survival. The RNA Integrity Number (RIN) score of 0 through 10 (perfect quality) is the gold standard for evaluating the integrity of the RNAFull paper Login or join for free to view the full paper.
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Discussion
Discussion
4 years ago
Author:
Paul G. Macon
RNA isolation from tissue
How do I extract RNA from animal tissue without using liquid nitrogen? I tried the RNA extraction by using the TRIzol reagent and I homogenize the tissue using polytron homogenizer at room temperature for 30secs is this correct?
Discussion
5 years ago
Author:
Aaron Stege
Problem in phase separation after using serum/plasma kit
I used a serum/plasma kit for my serum samples. After the phase separation the samples should have 3 phases: a colourless aqueous phase, a white interphase and a red organic phase. However, in some of my samples there was no aqueous phase unless I wait for an extended period of time. How can I circumvent this problem?
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Manufacturer protocol
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