RNeasy Mini Kit

RNA isolation / purification Bacteria - Gram positive Clostridum botulinum

Experiment
RNA isolation / purification Bacteria - Gram positive Clostridum botulinum
Product
RNeasy Mini Kit from Qiagen
Manufacturer
Qiagen

Protocol tips

Protocol tips
To yield high RNA, give the column an extra wash with both RW1 and RPE buffers (3 total washes with each buffer).
Downstream tips
"Include DNAse treatment for 15-20min.

Ensure EtOH is completely evaporated off of the column prior to elution. Adjust time from 1min to 5 min at 60`C.

Use water to elute the RNA that is warmed to ~60`C"

Publication protocol

Total RNA from C. botulinum ATCC 3502 and the two mutants was isolated using the RNeasy Mini Kit (Qiagen) as described [31]. Residual DNA was removed sequentially with RNase-free DNase set (Qiagen) and the DNA-free Kit (Ambion) according to the manufacturers' instructions. The RNA was dissolved in 50 µl of nuclease-free water (Sigma-Aldrich) and its concentration was determined using the NanoDrop ND1000 spectrophotometer (NanoDrop Technologies). The integrity of RNA was confirmed with the Agilent Technologies 2100 Bioanalyzer.

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Discussion

Discussion

4 years ago

Author: Paul G. Macon United States

Some help with RNA isolation using Trizol

Hello! I used Trizol to extract total RNA from in-vitro cultured bacteria (1 X 10^8 cells). After phase separation, I mixed ~0.4 ml of the upper phase which contains RNA with 0.5 mL cold isopropanol. However, the amount of RNA when measured in Nanodrop was very low. In addition, the ratio between 260 and 230 was around 0.1 to 0.5. Is there a chance that my sample was contaminated by the Trizol reagent? When I collected the aqueous phase I made sure to not touch the lower phase. What should I do?

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Papers

Check out relevant papers found by Labettor's AI that are relevant for performing RNA isolation / purification Bacteria - Gram positive Clostridum botulinum using RNeasy Mini Kit from Qiagen.

Paper title
Two-Component Signal Transduction System CBO0787/CBO0786 Represses Transcription from Botulinum Neurotoxin Promoters in ATCC 3502
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Manufacturer protocol

Download the product protocol from Qiagen for RNeasy Mini Kit below.

Download PDF Download manufacturer protocol

Videos

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