5R-PLEX KIT

General description

The 16S ribosomal RNA gene (16S rRNA) is a common bacterial marker, used as a target for microbial community profiling in microbiome metagenomic studies. The gene is composed of nine variable regions (V1-V9) interspersed between conserved regions. The most common 16S rRNA NGS assays target one or two regions using a single set of primers (e.g., V3-V4). However, this approach usually results in a poor bacterial detection and classification rate when the DNA input is of low-quality or when applying it on extremely low biomass samples.  The 5R-PLEX kit offers a complete solution to challenging biological samples where the standard 16S sequencing protocols fail to perform.   Sample types include:

• Formalin-Fixed, Paraffin-Embedded (FFPE) tissue
• Cancerous tumor tissue 
• Degraded or damaged DNA  
• Low biomass samples 
• Fossil-derived and ancient DNA

The 5R-PLEX kit was developed in collaboration with Dr. Ravid Straussman of the Weissmann Institute, based off a technique that was featured in his Science publication focused on the human tumor microbiome characterization (Science2020 May 29;368(6494):973-980. doi: 10.1126/science.aay9189) . The kit targets five short variable regions along the 16S rRNA gene (V2, V3, V5, V6, V8) that are co-amplified in a multiplexed PCR within a single tube.   After sequencing, the output data is uploaded to our M-CAMPTM platform (Microbiome Computational Analysis for Multi-omics Profiling) and analyzed using the unique 5R-PLEX algorithm[2]. This new analysis approach computationally combines the amplified regions to reconstruct one coherent high-resolution microbial profile. Read more about our kit here.

Features and Benefits

• The 5R-PLEX Kit provides high profile sequencing from degraded, damaged or low biomass samples
• Multiplexed NGS assay targets 5 variable regions of the 16S rRNA gene in a single primer pool
• Includes positive control for trouble shooting experiments.
• The kit can detect as low as 1pg of highly degraded bacterial DNA with an average fragment size of 260bp.
• Kit includes low bioburden reagents
• Includes exclusive access to M-CAMP^™ platform featuring an innovative algorithm designed to reconstruct a single coherent microbial profiling and perform comprehensive analysis.

Components

• 5R-PLEX PCR1 Primer mix- 25 μL
• 5R-PLEX PCR2 Forward Primer mix- 25 μL
• 5R-PLEX index plate 96-plate
• Water, microbial DNA-free 10X- 1.5 mL
• HF DNA Polymerase- 0.1 mL
• 5X HF buffer 2 mL
• dNTP′s- 0.2 mL
• Elution Buffer (EB), microbial DNA-free- 8 mL
• 5R-PLEX Positive Control (10 ng/μL)- 30 μL

Storage and Stability

The shelf life of all reagents provided in the kit is 12 months when stored properly. Store all components at -20 °C.

Other Notes

Kit component details can be found in the User guide.

Legal Information

M-Camp is a trademark of Merck KGaA, Darmstadt, Germany