Isolating RNA from the Soil
从土壤中分离RNA
发布: 2013年09月20日第3卷第18期 DOI: 10.21769/BioProtoc.903 浏览次数: 11796
评审: Anonymous reviewer(s)
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参见作者原研究论文
The authors used this protocol in:
Feb 2013
Abstract
Next generation sequencing has allowed for the analysis and ability to identify the microbial communities present in the environment. While DNA extraction from environments (such as soil) have provided a wealth of knowledge regarding microbial communities there are drawbacks that one encounters when using DNA as opposed to RNA. RNA allows for the determination of the identity of the microbes that are active and present at a particular time point and thus gives a clear picture of what these microbes are actually doing at a specific point in time and under a set of conditions. Extracting RNA from soil is challenging due to the inherent inhibitors present in the soil such as humic acids. Here we describe modifications to the MoBio RNA PowerSoilTM total RNA isolation kit to reproducively extract total RNA from the soil.
Materials and Reagents
- Fresh soil or soil stored at -80 °C
- RNA PowerSoilTM Total RNA Isolation Kit (MO BIO Laboratories, catalog number: 12866-25 )
- Phenol/Chloroform/Isoamyl Alcohol 25:24:1 (pH 6.7) (Thermo Fisher Scientific, catalog number: BP1752l-400 )
- Diethyl pyrocarbonate (DEPC) (Sigma-Aldrich, catalog number: D5758-50mL )
- RNase AWAY® (VWR International, catalog number: 72830-022 )
- Disposable Gloves
- Ice
- DEPC water (see Recipes) or nuclease-free water
Equipment
- Fisher Vortex Genie 2 (VWR International, model: 12-812 )
- Centrifuge (Sorvall® SuperT21 rotor SL-50T) (for 15 ml centrifuge tubes)
- Microcentrifuge (VWR International, model: Galaxy 16 )
- Shaker (New Brunswick Scientific, model: G-33 )
- Incubator set at 45 °C
- 5 ml Borosilicated Glass pipette (VWR International, model: 53281-818 )
- 1,000 μl pipetman
- 200 μl pipetman
- 2.5 μl pipetman
- -20 °C Freezer
- 4 °C Incubator
- Microcentrifuge Tube Rack (VWR International, model: CBGTR-080 )
- 15 ml falcon tube holder
- Laboratory labeling tape (VWR International, catalog number: 89097-902 )
- Spectrophotometer (e.g. Thermo Scientific, model: NanoDrop ND-1000 )
Procedure
文章信息
版权信息
© 2013 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Chaparro, J. M. and Vivanco, J. M. (2013). Isolating RNA from the Soil. Bio-protocol 3(18): e903. DOI: 10.21769/BioProtoc.903.
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分类
微生物学 > 微生物遗传学 > RNA
分子生物学 > RNA > RNA 提取
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