MICROBEnrich™ Kit
MICROB<i>Enrich</i>&trade; Kit
Invitrogen™

MICROBEnrich™ Kit

For the separation and isolation of bacterial RNA from mixed samples. The Ambion™ kit includes sufficient reagents for depleting 500Read more
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Catalog NumberQuantity
AM190120 preps
Catalog number AM1901
Price (USD)
808.00
Each
Add to cart
Quantity:
20 preps
Price (USD)
808.00
Each
Add to cart
For the separation and isolation of bacterial RNA from mixed samples. The Ambion™ kit includes sufficient reagents for depleting 500 μg of host cell RNA (20 reactions x 25 μg RNA) from a complex host-bacterial RNA mixture. Gene expression analysis following host-bacterial interactions is revolutionizing our understanding of the host response to bacterial infection. Numerous studies have utilized microarray analysis to follow host cell transcriptome alterations in response to interactions with bacteria. However, because no method has existed to isolate bacterial RNA away from host cell RNA, the corresponding analyses of bacterial transcriptomes has been impossible. The MICROBEnrich™ Kit enables whole genome microarray expression analyses of bacterial RNA after host-bacterial interactions.

Start With Any Total RNA Sample
For microarray analysis and other sensitive applications, a DNase treatment is recommended to remove contaminating genomic DNA from the total RNA prior to the MICROBEnrich™ Kit procedure.

Remove >90% of Mammalian RNA from Complex Total RNA Mixtures
The MICROBEnrichKituses hybridization capture technology (patent pending) to remove human, mouse, and rat RNA (both mRNA and rRNA) from complex host-bacterial RNA populations, leaving behind enriched microbial total RNA. In the first step of the procedure, host-bacterial total RNA is incubated with a mixture of capture oligonucleotides that bind the mammalian 18S and 28S rRNAs and polyadenylated RNAs. Next, the rRNA/oligo nucleotide hybrids and all polyadenylated mRNAs are removed from the mixture with oligonucleotide-derivatized magnetic beads. After magnetic separation, the bacterial RNA remaining in the supernatant and can be precipitated with ethanol. An Agilent 2100 bioanalyzer electropherogram (see Figure) shows nearly complete removal (>95%) of the host 18S and 28S rRNAs from a mixed sample. Bacterial mRNAs purified with the MICROBEnrich™ Kit are better templates than total RNA for downstream applications and therefore deliver dramatic increases in sensitivity when used in applications such as array analysis (see Figure).

The MICROBEnrich™ Kit Will Work With Any Bacterial Species
The MICROBEnrich Kit was optimized using E. coli RNA in a background of human, mouse, and rat RNA. However, the kit should perform well with RNA from any bacterial species. Although many mammalian host species may be compatible with the MICROBEnrichKit, only human, mouse, and rat rRNA sequences were used for the design and optimization of the capture oligonucleotides.

Accessory Products:
A magnetic stand is required for use this kit. They are available in several formats, including the Single Place Magnetic Stand (SKU #AM10026), 6 Tube Magnetic Stand (SKU #AM10055), and 96-well Magnetic Stands (SKU #AM10027 and #AM10050). The RiboPure™-Bacteria Kit (SKU #AM1925) is ideal for the purification of host-bacterial cell total RNA. The TURBO™ DNA-free™ or DNA-free™ DNase Treatment and Removal Reagents (SKU #AM1907 and AM1906, respectively) are recommended for microarray analysis and other applications where the removal of contaminating genomic DNA is needed.After the MICROBEnrich™ procedure, bacterial mRNA can be purified from the total RNA using the MICROBExpress™ Bacterial mRNA Enrichment Kit (SKU #AM1905). This technology enables >95% removal of prokaryotic 16S and 23S rRNAs.
For Research Use Only. Not for use in diagnostic procedures.
Specifications
Final Product TypeTotal RNA (Bacterial)
For Use With (Application)Microarray Analysis
High-throughput CompatibilityNot High-throughput Compatible (Manual)
No. of Reactions20 Preps
Product LineAmbion™, MICROBEnrich
Quantity20 preps
Isolation TechnologyHybridization Capture, Magnetic Bead
Sample TypeBacteria, RNA, Total RNA (Mammalian Host-Bacterial RNA Mixture)
TargetTotal RNA
Unit SizeEach
Contents & Storage
Store Control RNA, Capture Oligo Mix, Glycogen, and 3M NaOAc at -20°C.

Store Oligo MagBeads, Binding Buffer, Wash Solution, Regeneration Solution 1, Regeneration Solution 2, and Resuspension Solution at 4°C.

Store Nuclease-free Water, 1.5 mL tubes and 2 mL collection tubes at room temperature.

Frequently asked questions (FAQs)

I am using the MICROBEnrich Kit to separate bacterial RNA from mouse RNA but am having problems with RNA degradation. Can you offer some tips?

Here are possible causes and solutions:

1. Input total RNA is degraded: to see whether the RNA used in the MICROBEnrich procedure was degraded from the start, evaluate a 0.5-5 µg sample of the input RNA mixture on a denaturing agarose gel (see section III.C starting on page 15 of the manual [https://tools.thermofisher.com/content/sfs/manuals/cms_057050.pdf]). Total RNA should produce rRNA bands that appear sharp and well-defined (Figure 3 on page 16). In high-quality RNA samples, the 28S rRNA band will be 1.5-2 fold brighter than the 18S rRNA band.
2. Practice RNase-free technique: all of the typical precautions against RNase contamination should be observed. Gloves should be worn at all times and changed frequently to avoid the introduction of RNases. Bags containing the centrifuge tubes and the solution tubes and bottles should be kept closed when they are not in use to avoid contamination with dust. Any tubes or solutions that will contact the RNA (that are not supplied with the kit) should be free of RNases.
3. Degradation caused by the heat denaturation step: RNA degradation can occur at elevated temperatures when there are divalent cations in the solution. If your RNA solution does not contain at least 1 mM EDTA, contaminating divalent cations could cause RNA hydrolysis during the heat denaturation step (step II.B.3 on page 8).