Catalog No. : MN-740140.20
NucleoBond RNA Soil
NucleoBond RNA Soil for the isolation of RNA from soil samples, NucleoBond Columns, buffers
Can be combined with 740141.20 for parallel DNA isolation
RNA purification from soil samples for metagenomic analysis
- Anion exchange technology to optimize RNA yield and purity - suitable for metagenomic studies
- Combination of mechanical homogenization and chemical lysis allows processing of large sample amounts
- Parallel preparation of RNA and DNA in one hour
|NucleoBond RNA Soil||
NucleoBond® RNA Soil Columns, NucleoSpin® Bead Tubes Type A, NucleoSpin® Finisher, buffers
- RNA isolation from soil for metagenomic analysis
- Typical downstream applications: RNA sequencing, real-time RT-PCR, Northern blotting, primer extension, array technology, RNase protection assays
* Kits to be used for research purposes only
The NucleoBond® RNA Soil Kit is designed for the purification of RNA from soil and sediment samples. In combination with the DNA Set for NucleoBond® RNA Soil, RNA and DNA can be isolated from the same soil sample. Thus, purified nucleic acids are suitable e.g., for metagenomic studies, where the genome and transcriptome are analyzed from the same sample.
Most soils contain relatively low amounts of RNA, compared to DNA. Therefore, the sample input needs to be increased in order to gain acceptable RNA yields. The standard protocol of this kit uses four NucleoSpin® Bead Tubes Type A that can be filled with 250–500 mg of one soil sample each, resulting in a total sample amount of 1–2 g. This sample input amount will usually result in a RNA yield in the range from 1–10 μg. If working with four bead tubes per sample is too cumbersome, the content of four bead tubes can be combined into one 15 mL tube. This tube can be filled with 1–2 g of soil sample and four times the volume of Buffer E1, PCI and, if applicable, Buffer OPT. On the other hand, if a specific soil contains high amounts of RNA, the number of NucleoSpin® Bead Tubes Type A can also be reduced.
Organisms in a soil sample are lysed by bead beating in the presence of Lysis Buffer E1 and Phenol:Chloroform:Isoamylalcohol. Buffer OPT reduces the adsorption of nucleic acids to clay and mineralic soil components, but will also increase the contamination with humic acids if present. Performance cannot be predicted and depends on the soil composition, but as a rule of thumb a sample with a high content of organic and humic components, e.g., forest soil, should be lysed without Buffer OPT while predominantly mineralic soils, e.g., river sediments and clay, should be lysed with Buffer OPT.
Unlysed components are sedimented by centrifugation. The supernatants of the four bead tubes are transferred and combined into one fresh 15 mL tube and mixed with Binding Buffer E2. The incubation and centrifugation steps that follow can be used to prepare the NucleoBond® RNA Columns. The columns including filters are fixed with the supplied Plastic Washers on top of a 50 mL tube or laboratory flask or any NucleoBond® Rack. Filters and columns are equilibrated by Buffer EQU to pre-wet the filters and to prepare the anion exchange chromatography columns.
Once the columns are equilibrated, the clear supernatant of the centrifuged samples is loaded onto the filters in the purification columns. Undissolved particles and some of thesoluble macro molecules will be held back by the filters.
Wash Buffer E3 is used to flush the filter columns and to wash the nucleic acids from the filters onto the column matrix. Nucleic acids and soluble polyanionic molecules including some fraction of humic substances (if present) will bind to the anion exchange surface. A brown layer might be visible on top of the silica matrix. After the washing step the filter is removed and the anion exchange column is washed with Buffer E4 which removes contaminants.
RNA is eluted by Buffer ERNA. If isolation of RNA and DNA is desired, DNA is eluted by Buffer EDNA, contained in the DNA Elution Set (to be ordered separately, see section 5), from the same column. Isopropanol is added to the eluates and the mixture is bound to NucleoSpin® Finisher Columns by centrifugation. The columns are washed with Buffer E5, dried and eluted inRNase-free H2O.
NucleoBond® RNA Soil procedure
High RNA yields with NucleoBond® RNA Soil
Different soil samples (clay, shore soil, forest soil) were purified in duplicates according to the standard procedure. For comparison, the samples were applied to a competitor kit from Q. RNA was eluted in 100 µL and determined photometrically. 15 µL of eluates were used for gel electrophoresis (1 % TAE). NucleoBond® RNA Soil convinced due to the high RNA yield.
Amplifiable RNA for perfect results with NucleoBond® RNA Soil kit
Duplicates of different soil samples (clay, shore soil, forest soil) were purified in duplicates according to the standard procedure from MN and Q. 40 µL of eluate was applied to the RT-PCR (amplicon: 466 bp). All MN samples showed low CP values compared with Q samples.
NucleoBond® RNA Soil enables efficient parallel DNA isolation
Different soil samples (clay, shore soil, forest soil) were purified in duplicates according to the standard procedure. Samples were also purified according to a standard procedure of a competitor kit from Q. DNA was eluted in 100 µL and measured photometrically. 15 µL of eluates were applied to 1 % TAE gel for electrophoresis analysis. High DNA yields can be purified using NucleoBond® RNA Soil with the DNA Set for NucleoBond® RNA Soil.