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AllPrep DNA/mRNA Nano Kit

For the simultaneous purification of genomic DNA (gDNA) and messenger RNA (mRNA) from low-biomass samples
  • Simultaneous gDNA and mRNA isolation
  • Maximal yields from minimal cell quantities
  • High-molecular-weight, sequencing-ready gDNA
  • Messenger RNA suitable for single-cell RNA-seq application
  • Magnetic-bead-based system

Collect genetic and transcriptomic information from CTCs and rare cells, with minimal cell quantities, by isolating gDNA and mRNA from the same sample, at the same time.

The AllPrep DNA/mRNA Nano Kit uses a two-step magnetic-bead-based method that lets you maximize mRNA capture and isolate high-molecular-weight DNA for multiomics studies.

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Cat No./ID: 399911
AdnaMag-S
$728.00
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For 8 tubes, 1.5 ml
Cat No./ID: 80272
AllPrep DNA/mRNA Nano (12)
$599.00
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For 12 preps: Oligo-dT magnetic beads, DNA-isolation beads, collection tubes, RNase-free water and buffers

The AllPrep DNA/mRNA Nano Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

Product Details

9
Simultaneous mRNA and gDNA purification.

Six samples containing AllPrep DNA/mRNA Nano lysis buffer were each spiked with 5 MCF-7 cells and 5 LNCaP95 cells. Messenger RNA and genomic DNA were isolated using the AllPrep DNA/mRNA Nano Kit. Messenger RNA was analyzed using the AdnaTest BreastCancerDetect Kit (GA733-2 expression profile), and matched genomic DNA yield (H91; human target, small autosomal, 91 bp) was determined using the Investigator Quantiplex Pro RGQ Kit.

A range of 7.9–11.9 ng/µl (median: 10 ng/µl) of mRNA was isolated, and gDNA was detected within 28.3–29.6 cycles (median Cq: 28.8).

9
Minimal amount of cells required.

AF=allele frequency.

Increasing numbers of MCF-7 cells were spiked into 5 ml of healthy donor (HD) blood and subsequently recovered using the AdnaTest CTC-Select procedure. Genomic DNA was isolated using AllPrep DNA/mRNA Nano. Pyrosequencing for PIK3CA E545K was done using the PyroMark Q48 Autoprep.

MCF-7 PIK3CA hotspot mutation was detected by Pyrosequencing down to 5 MCF-7 cells in a background of 500–1000 co-isolated wild-type leukocytes.

9
Maximal gDNA and mRNA yields.

Six samples that contained AllPrep DNA/mRNA Nano lysis buffer were each spiked with 5 MCF-7 cells and 5 LNCaP95 cells and further processed according to the AllPrep DNA/mRNA Nano Kit protocol. Subsequently, mRNA (Ga733-2 and Her2 expression profiles) was analyzed using the AdnaTest BreastCancerDetect Kit; matched gDNA yield (H353 and H91) was determined by PCR using the Investigator Quantiplex Pro RGQ Kit.

Genomic DNA and messenger RNA yields exceeded all other competitor procedures significantly.

9
High-molecular-weight genomic DNA.

Genomic DNA was isolated from various numbers of MCF-7 cells and analyzed with an automated electrophoresis instrument.

Genomic DNA isolated with the AllPrep DNA/mRNA Nano Kit has high molecular weights of 60,000 bp or more.

9
Highly suitable for long-range PCR.

Genomic DNA was purified from ≤2000 cells and amplified using the UltraRun LongRange PCR Kit.

A 4 kb amplicon from P53 was amplified from as few as 20 cells.

The resulting PCR fragments are highly suitable for a variety of applications, such as cloning, genome mapping and sequencing.

9
mRNA for single-cell RNA-seq applications.

Messenger RNA from single cells was purified using the AllPrep mRNA/DNA Nano procedure. RNA sequencing was done with the QIAseq UPX 3' Transcriptome Kit for high-throughput gene-expression analysis for single-cell CTC analysis of metastatic breast cancer (Breast CTC) samples and primary ovarian cancer (Ovary CTC) samples.

The UPX gene-expression profiles differentiate single cells from Breast CTC, Ovary CTC, and healthy donor controls.

9
Sequencing-ready gDNA from low cell numbers.

VAF: variant allele frequency.

One hundred sixty-six cells from each of the different cell lines were spiked into a healthy donor background. Cells were recovered using the AdnaTest CTC-Select Kit. Messenger RNA and genomic DNA were isolated using the AllPrep DNA/mRNA Nano Kit.

Isolated DNA was used for library preparation, and NGS was performed using the QIAseq Human Solid Tumor Panel.

All variants described for the selected cell lines were detected with the expected allele frequency. None of them were found in the healthy donor control.

9
Magnetic-bead-based system.

The AllPrep DNA/mRNA Nano Kit uses a two-step magnetic-bead-based isolation process. This maximizes mRNA capture efficiency and allows high-molecular-weight gDNA isolation for multiomics studies.

Performance

The AllPrep DNA/mRNA Nano Kit purifies high-quality gDNA and mRNA from the same cell (see figure “Simultaneous mRNA and gDNA purification”) or low-biomass input samples (see figures “Messenger RNA for single-cell RNA-seq applications”, “Sequencing-ready gDNA from low cell numbers”, and “Minimal amount of cells required”).

The kit’s simple and convenient procedure does not require the use of toxic substances such as phenol (see figure “Magnetic-bead-based system”).

Genomic DNA purified with the AllPrep mRNA/DNA Nano Kit procedure has an average length of 60–100 kb, depending on homogenization conditions (see figure “High-molecular-weight gDNA”). Compared to products from other suppliers, the AllPrep mRNA/DNA Nano Kit provides nucleic acids of significantly higher yields (see figure “Maximal DNA and mRNA yields”).

Principle

Unlike other procedures where the biological sample or purified total nucleic acids are divided into two before being processed separately, the AllPrep DNA/mRNA Nano Kit purifies gDNA and mRNA simultaneously from the same precious sample. It also allows the parallel processing of multiple samples in less than 75 minutes.

The kit is compatible with small amounts of low-biomass samples such as CTCs, rare cells and low numbers of cells of animal or human origin.

The use of toxic substances such as phenol and/or chloroform, and tedious methods such as alcohol precipitation, are replaced by safe and simple magnetic-bead-based workflows using a magnetic rack such as the AdnaMag-S.

Procedure

The AllPrep mRNA/DNA Nano Kit’s magnetic-bead-based system purifies mRNA using oligo-dT magnetic beads, while gDNA is purified using DNA-affinity beads.

Applications

Genomic DNA purified with the AllPrep DNA/mRNA procedure has a high molecular weight of up to 100 kb. This purified DNA is suitable for any downstream application, such as PCR, multiplex PCR, dPCR, pyrosequencing or NGS.

The purified mRNA is of high quality and can be used in any application, such as RT-PCR, real-time RT-PCR, or cDNA generation, including the AdnaTest procedure for molecular characterization of CTCs.

Specifications

Features
Specifications
Applications Transcriptome and genome profiling, real-time PCR, cDNA synthesis, NGS, RNA seq
Elution volume 25 µl
Format Magnetic-bead-based system
Main sample type Low-biomass samples (rare cells, CTCs, fetal cells, stem cells, T-cells)
Processing Manual (magnetic rack)
Product use Purification of full-length messenger RNA and high-molecular-weight genomic DNA
Sample amount Down to 1 cell
Technology Magnetic beads
Time per run or per prep 75 min per prep

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