Archer® RNA Reagent Kits v2


Universal RNA Reagent Kit v2 for Illumina and Ion Torrent platforms

The Archer® Universal RNA Reagent Kit v2 is an integral part of the Archer FusionPlex® system, which includes Molecular Barcode Adapters and a target-specific FusionPlex assay.

The Universal RNA Reagent Kit v2 contains all of the enzymes and buffers needed to create a target-specific library for next-generation sequencing and now includes a sample input quality assay and an improved DNA polymerase.

Archer Universal RNA Reagents are now included in all Archer FusionPlex kits.

Archer Universal RNA Reagents are now included in all Archer FusionPlex kits. Effective February 5th, 2016 all Archer FusionPlex Kits will include all buffers, enzymes, reagents and previously provided separately in Archer Universal Reagent Kits, v2. PreSeq QC Assays will also be combined into the new kit format. Archer MBC Adapters will continue to be provided separately to preserve flexibility and index diversity.

Please note that while the product number for the kits have changed, the formulations, component part numbers and shipping conditions have not. Individual assay components will be provided with lot numbers and expiration dates. Furthermore, there will be no increase in the price of the historical components that make up the new kit format.


Have any questions about the change in format? Please contact us at tech@archerdx.com or chat with an online representative



User-Friendly Workflow


 


RNA Workflow

The workflow is a series of sequential transfers of input nucleic acid into tubes that contain lyophilized pellets of all of the enzymes and buffers required for each reaction step.

  • Random priming - random hexamers hybridize to the RNA to generate random start sites for cDNA synthesis
  • First strand synthesis - reverse transcriptase binds to the primed RNA and synthesizes a single-stranded cDNA
  • PreSeq™ RNA QC assay - verify sample integrity prior to sequencing by analyzing a small aliquot of the cDNA by qPCR
  • Second strand synthesis - remaining RNA in the sample is digested and polymerase synthesizes the second cDNA strand
  • End repair/dA-tailing - the ends of the double-stranded cDNAs are blunted, phosphorylated and adenylated to prepare for adapter ligation
  • Adapter ligation - MBC adapters are ligated to the end-polished cDNA
  • First PCR - target regions are selectively amplified using gene-specific primer 1 (GSP1)
  • Second PCR - target regions are further enriched with a nested gene-specific primer 2 (GSP2)

PreSeq RNA QC Assay


PreSeq RNA QC Assay

The Archer PreSeq RNA QC Assay is used to identify samples that would likely fail sequencing due to insufficient quality of RNA in a sample. RNA samples that have been significantly degraded through formalin fixation, aging or poor handling can be ruled out of the testing queue to save time and money.

Existing methods to measure RNA quantity do not adequately predict the quality or amount of amplifiable RNA in the sample, which ultimately determines successful sequencing library generation. PreSeq overcomes this shortfall by priming and amplifying a ubiquitously expressed control gene of known segment length by qPCR. Samples where the integrity of RNA has been compromised can be identified prior to sequencing. PreSeq can also be used post-analysis to troubleshoot failed samples and unexpected negative fusion calls. While there is a strong correlation between qPCR results and predicted fusion calling success, laboratories should empirically determine their own threshold for predicted sample failure based upon their needs and testing methods.

The PreSeq RNA QC assay is incorporated into the Archer Universal RNA Reagent Kit v2 protocol between First Strand cDNA Synthesis and Second Strand cDNA Synthesis. Data should be analyzed prior to First PCR so that other samples can be substituted in the case of sample failure. For this reason, additional sets of Random Priming, First Strand cDNA Synthesis and Second Strand cDNA Synthesis reagents are included with the kit.


Lyophilized Reagents


FusionPlex-lyophilized-reagents


The reagents are lyophilized, and single reactions are aliquoted into 8-tube strips, eliminating the need to make master mixes and minimizing the risk of sample cross-contamination. The pouch labels and tubes for each step are also color coded for easy identification.


Modular Assay Design

The Archer FusionPlex system is a flexible method to generate target-specific libraries for next-generation sequencing. The modular nature of the system ensures platform compatibility and accurate library preparation. To generate a library for sequencing, you will need one of each of the following:

  1. An application-specific FusionPlex Assay
  2. Platform-specific Molecular Barcode (MBC) adapters
  3. A platform-specific Universal RNA Reagent Kit v2

Universal Reagent Kit

Universal-RNA-Kit-v2



Kit contents

The Universal RNA Reagent Kit v2 contains reagents for 8 reactions and include the PreSeq RNA QC Assay primer mix to perform qPCR after first strand cDNA synthesis.

Random Priming reagent two 8-tube strips
First Strand cDNA Synthesis reagent two 8-tube strips
PreSeq RNA QC Assay 10X VCP Primer Mix 50µL
Second Strand cDNA Synthesis reagent two 8-tube strips
End Repair/dA-Tailing reagent one 8-tube strip
Adapter Ligation reagent one 8-tube strip
First PCR reagent one 8-tube strip
Second PCR reagent one 8-tube strip

The kits also include 2 8-tube strips of the first 3 reagents to give you the greatest flexibility in confirming sample quality prior to sequencing. For example, screen up to 16 samples using the PreSeq RNA QC Assay to identify 8 quality samples to use in downstream library preparation. And while performing the QC assay, proceed to second strand cDNA synthesis with up to 16 samples to ensure sample stability.


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How to contact us

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Boulder, CO 80301

Phone

Phone: (877) 771 1093

Phone: (303) 357 9001

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For Research Use Only. Not for use in diagnostic procedures. For Research Use Only. Not for use in diagnostic procedures.