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ThruplexPlasma-seq

ThruPLEX Plasma-SEQ Kit

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ThruPLEX® Plasma-seq is powered by ThruPLEX chemistry to generate high performance NGS libraries from cell-free DNA isolated from plasma.

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Product Description

ThruPLEX® Plasma-seq is powered by ThruPLEX chemistry to generate high performance NGS libraries from cell-free DNA isolated from plasma.

The chemistry is optimized specifically for cell-free DNA to maximize the library complexity and to preserve the GC representation of the input DNA. Libraries can be used for CNV analysis, whole genome sequencing applications or enriched using your custom panels or leading target enrichment platforms, such as Agilent SureSelect® and Roche NimbleGen SeqCap® EZ.

Libraries prepared with ThruPLEX Plasma-seq provide added sensitivity to better understand genetic variations found within the plasma samples

Advantages

  • Designed for cell-free DNA: newly formulated repair and ligation reagents with optimized protocols
  • High performance NGS libraries: high diversity with broad and reproducible GC coverage
  • Variable sample input: <1 ng to 30 ng of cell-free DNA
  • Fast and simple workflow: 3 steps in a single tube or well in 2 hours with no purification or sample transfer steps
  • Research applications: libraries from samples where results require high sensitivity including liquid biopsies, ctDNA, and targeted sequencing
  • Automation-friendly: Beckman®FXPWorkstation

Specification

Guaranteed for 9 months at -20°C in a constant temperature freezer.

Store at –20 °C.

ThruPLEX Plasma-seq contains all necessary reagents for preparing indexed Illumina NGS libraries, including optimized Illumina-compatible adapters and indexing reagents.

Performance

Highest Diversity, Fewest Unmapped Reads from Cell-Free DNA

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ThruPLEX Plasma-seq Kit generated quality libraries with high diversity and a low number of duplicates and unmapped reads. Cell-free DNA was extracted from 3 plasma samples, and libraries were prepared at the amounts indicated as measured by Qubit®. The amount of mononucleosomal DNA in each sample as measured by the Bioanalyzer® corresponded to 0.09 ng, 0.62 ng, and 15.44 ng. Pooled libraries were sequenced on an Illumina NextSeq® 500 as a paired-end run with 17M to 25M reads per library, Duplication rates were calculated after down-sampling the data to 17 M reads per library.

Outstanding Target Enrichment Peformance

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ThruPLEX Plasma-seq Kit libraries were captured at high efficiency and generated data with deep coverage of the kinome for mutation detection. Libraries were prepared from 3 plasma samples at input amounts of 5 ng, 6.5 ng, and 10 ng in triplicate, and targeted sequencing was carried out on an Illumina MiSeq using samples enriched with the ClearSeq Human DNA Kinome Panel for SureSelectXT2. On average, 5M reads were generated per library. Selected bases were successfully captured bases that were in or within 250 bp of the baits.

Reproducible, Unbiased GC Coverage

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ThruPLEX Plasma-seq Kit provided the most reproducible and unbiased GC coverage across the human genome. ThruPLEX libraries showed minimal variability across 9 individual plasma samples tested. Libraries were prepared from cell-free DNA isolated from 1 mL of plasma samples and sequenced on an Illumina NextSeq 500. Four separate plasma samples were used to construct the NEBNext Ultra libraries.

Evaluated at Karolinska Institute and CRUK

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Illumina NGS libraries were prepared with 5 ng of cell-free DNA isolated from pooled plasma samples using ThruPLEX Plasma-seq Kit. Low-pass WGS was conducted on a HiSeq with approximately 12.5M reads per library. (Data courtesy of the Karolinska Institute, Sweden).

ThruPLEX Plasma-seq Kit Single-Tube Workflow

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Starting with 1 to 30 ng of cell-free DNA, ThruPLEX Plasma-seq Kit creates indexed libraries in 3 simple steps: end repair, adapter ligation, and high-fidelity library amplification. No purification or sample transfer steps are required. The streamlined workflow is performed in 2 hours in a single tube or well, preventing sample loss and enhancing positive sample identification.

ThruPLEX Plasma-seq Kit Technology

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ThruPLEX Plasma-seq Kit technology is a 3-step reaction that is optimized for cell-free DNA. Cell-free DNA (1 ng to 30 ng) is first repaired in a highly efficient process. Background is reduced using double-stranded adapters with no single-stranded tails. Blunt-end ligation occurs with high-efficiency. Blocked 5’ ends reduce adapter-adapter ligation. Background is further reduced by destroying unused adapters after ligation. A high-fidelity amplification completes the reaction to generate indexed Illumina libraries.

FAQ

1. What types of samples and inputs can be used with ThruPLEX Plasma-seq?
2.What are the major technical differences between ThruPLEX DNA-seq Kit and the new ThruPLEX Plasma-seq Kit?
3.What are the protocol differences between the ThruPLEX Plasma-seq and ThruPLEX DNA-seq?
4.Can the reagents from ThruPLEX Plasma-seq be interchanged with the reagents from ThruPLEX DNA-seq?
5.What type of indexes or barcodes do you have available?
6.What recommendations do you have for isolating the cell-free DNA from plasma?
7.What kind of collection tube should I use to draw blood from subjects?
8.How should I prepare plasma from blood before isolating the cell-free DNA?
9.Can cell-free DNA isolated from urine or other biofluids be used with ThruPLEX Plasma-seq Kit?
10.Is ThruPLEX Plasma-seq Kit compatible with any target enrichment systems?

Download

ThruPLEX Plasma-seq Kit Instruction Manual

Blood Collection and Plasma Preparation Protocol

ThruPLEX Plasma-seq Product Sheet

ThruPLEX Plasma-seq Quick Protocol

ThruPLEX Plasma-seq Kit Index Guide

ThruPLEX Plasma-seq Index Sequences 12S (CSV)

ThruPLEX Plasma-seq Index Sequences 48S (CSV)

ThruPLEX Plasma-seq SDS

Animal-Origin Statement

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