BioLiqX cf-RNA Isolation QC Kit

Description

BioLiqX cf-RNA Isolation QC Kit for checking cf-RNA isolation efficiency

The BioLiqX cf-RNA Isolation QC Kit contains all reagents and primers required for the detection of four “house-keeping” miRNAs hsa-miR-16, hsa-miR-21 and hsa-miR-451a, and cel-miR-39 in any RNA sample using RT-qPCR. The kit also contains synthetic cel-miR-39 for spike-in assessment of small cf-RNA isolation between samples as well as normalization of miRNA expression in biological fluids after downstream qRT-PCR.

The key features of the method are highly efficient polynucleotide tailing and simultaneous reverse transcription of all short RNA species in a sample which are followed by real-time PCR amplification. The tailing and reverse transcription (TaRT) mix included in the kit is an optimized composition of enzymes, nucleotides, mineral salts, and RT primers that secures an efficient conversion of highly diluted miRNA molecules into cDNA in a single reaction. In the second step, the cDNAs serve as templates for ultra-sensitive and low-background real-time PCR reactions with Green DNA Dye (a SYBR® Green analog) and miRNA-specific primers. Due to its simplicity, the procedure secures low variation between technical replicates, short hands-on time, and easiness of automation. High specificity and sensitivity of qPCR are achieved by optimal concentration of nucleotides, mineral salts, Green DNA Dye and Hot Start Taq DNA Polymerase in the reaction. All reactions can be set up at the room temperature without the risk of non-specific amplification. The BioLiqX cf-RNA Isolation QC kit is ideal for monitoring the RNA isolation efficiency between samples and has a similar sensitivity as compared to analogous preamplification-free miRNA assays.

Workflow of the BioLiqX cf-RNA Isolation QC Kit

In the first step, polyA-tails are added to 3’-OH termini of all RNAs and cDNA is synthesized simultaneously using an anchored dT-rich primer with custom 5’-terminal sequence. Subsequently, cDNA is amplified using miRNA-specific forward and reverse primers during qPCR reaction, while Green DNA Dye (a SYBR® Green analog) is used for the detection and quantification of dsDNA amplicons in real-time

Kit Contents

The kit contains ready-to-use TaRT Mix for tailing and reverse transcription, qPCR Mix for Real-Time PCR reaction, as well as four vials with miRNA-specific primers: hsa-miR-16 PM, hsa-miR-21 PM, hsa-miR-451a, and cel-miR-39 PM. The kit also includes synthetic cel-miR-39 spike-in control.
By default, the kit is supplied for 24 reactions.

Kit Customization

Any kit sizes larger than 24 rxns can be manufactured for you –
please contact us for a quote!

Related products:

For detecting low copy number circulating miRNAs in biological fluids Heidelberg Biolabs have developed BioLiqX HS miRNA qRT-PCR Assays, based on the so-called “Capture and Amplification by Tailing and Ligation (CATL)” approach for cDNA synthesis and pre-amplification of short or fragmented RNA in the original sample before the final Real-Time PCR step.