Thermo Fisher Scientific Axiom Quick Reference

QUICK REFERENCE

Axiom™ 384HT and Mini 96 gDNA Sample Preparation

Pub. No. MAN0017719 Rev. B.0

Table 1 Genomic DNA sample input requirements

WARNING! Read the Safety Data Sheets (SDSs)

and follow the handling instructions. Wear appropriate
protective eyewear, clothing, and gloves. Safety Data
Sheets (SDSs) are available from thermofisher.com/
support.

Sample type

Human 8.7 µL 100 ng 11.5 ng/µL

Diploid plants
and animals

Volume per

well

8.7 µL 150 ng 17.2 ng/µL

Input mass per

well

gDNA

concentration

Introduction

Running the Axiom™ 2.0 Assay for 384 and mini 96-array format
samples requires the following sets of steps:

1. Preparation of genomic DNA, as described in this document.

2. Target preparation of the samples, as described in one of the
following quick reference documents:

• Axiom™ 2.0 Assay 384HT Array Format
Automated Workflow Quick Reference—Biomek™ FX
(Pub. No. 703270)

• Axiom™ 2.0 Assay 384HT Array Format Automated
Workflow Quick Reference—Biomek™ i7 Method V1.0
(Pub. No. MAN0017557)

• Axiom™ 2.0 Assay Mini 96‑Array Format Manual
Workflow Quick Reference (Pub. No. 703436)

3. Array processing, as described in GeneTitan™ MC Protocol

for Axiom™ 384HT Array Plate Processing Quick Reference

(Pub. No. MAN0017596).

IMPORTANT!

instructions. Read all the instructions in Chapter 2, Genomic DNA
preparation and requirements of the appropriate user guide for

more details on the protocol and sample requirements.

Axiom™ 2.0 Assay 384HT Array Format Automated Workflow

·

User Guide—Biomek™ FXP (Pub. No. 703268)
Axiom™ 2.0 Assay 384HT Array Format Automated Workflow

·

User Guide—Biomek™ i7 (Pub. No. MAN0017555)
Axiom™ 2.0 Assay Mini 96‑Array Format Manual Workflow User

·

Guide (Pub. No. 703434)

This document contains an abbreviated set of

P

Polyploid plants
and animals

8.7 µL 200 ng 23.0 ng/µL

IMPORTANT! Prepare your genomic DNA sample plate in

a clean room. The clean room should be separate from the
laboratory where the assay is performed and must be free of DNA
amplified in other procedures.

Note: Use at least one positive control DNA sample on each
plate. For human samples, Genomic DNA Standard (Ref 103)
(Cat. No. 951957) can serve as the control. For plant or
animal samples, use a genomic DNA sample that meets the
specifications that are mentioned above, is from the same species
that is represented on the array, and ideally has passed previously
in the Axiom™ Genotyping Solution. If no DNA control for your
human sample type is available, then the Genomic DNA Standard
(Ref 103) can serve as a positive control during target preparation.

Reagents required

Reagent

Genomic DNA Standard (Ref 103) (positive control for human
samples), Cat. No. 951957, –20°C

Reduced EDTA TE Buer (10 mM Tris-HCl pH 8.0, 0.1 mM EDTA)
Cat. No. 75793

Positive control gDNA (if genotyping non-human samples)

Requirements

Review this information before proceeding with the DNA
amplification for the automated target preparation.

The genomic DNA (gDNA) you process must meet the following
requirements:

• Starting DNA must be double-stranded for accurate
concentration determination.

• DNA must be of high purity and be free of DNA polymerase
inhibitors.

• DNA must not be degraded.

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

Equipment and consumables required

Quantity Item

As required Adhesive seals for plates

1 Ice bucket, filled with ice

1 each Pipettes: single channel P10 or P20

Optional: multichannel P10 or P20

• Thaw in a water bath:
a. Fill a small plastic dish with ultra-pure water (such

as Millipore™ water). Do not overfill. The level of
the water must not overflow when the sample
tubes or plates are placed in the bath.

b. Place the DNA samples in the water bath and

thaw for 30 minutes.

c. Wipe water o the sample plate before removing

the seal to avoid contamination of the samples.

As required Pipette tips

1 Plate, deepwell:

• For the Axiom™ 2.0 Assay 384HT Array Format
Automated Workflow
Axygen™ 384 well clear V-bottom 240 μL
polypropylene deep-well non-treated plate, sterile
(Cat. No. P-384-240SQ-C-S)

• For the Axiom™ 2.0 Assay Mini 96‑Array Format
Manual Workflow, one of the following:

– Eppendorf™ 96 deepwell, 2,000 μL (Cat. No.

951033481)

– Square 96 Deepwell Plate, 2 mL

1 Plate centrifuge

1 Microtiter plate fluorimeter

Quant-iT™ PicoGreen™ dsDNA Assay Kit (required only if
no concentration measurements available for samples)

1 Vortexer

[1]

This plate is part of the Axiom™ 384HT Consumables Kit for Biomek™ FXP (Cat.
No. 902288), and the Axiom™ 384HT Consumables Kit for Biomek™ i7 (Cat. No.

905424). If purchased directly from the supplier, a unique barcode must be added.

[2]

This plate is part of the Axiom™ 2.0 Assay Mini 96 Manual Target Preparation
Consumables Kit (Cat. No. 902986).

[3]

This plate is part of the Axiom™ 2.0 Assay Mini 96 Manual Target Preparation
Consumables Kit v2 (Cat. No. 952431).

(ABgene™ 96-well 2.2 mL Polypropylene Deepwell Storage Plate, Cat. No. AB0932 )

[2]

[1]

[3]

Prepare genomic DNA samples

1. Prepare gDNA control sample.

• If genotyping human samples, use the Genomic DNA
Standard (Ref 103) (Cat. No. 951957).

• If genotyping non-human samples:

– Select a sample to be the positive control.

Ensure that it is double-stranded, highly pure,
is free of DNA polymerase inhibitors, and is not
degraded. Ideally, this sample has shown passing
performance when used in the Axiom™ Genotyping
Assay.

– Dilute the sample to a working concentration.

For diploid plants and animals, dilute the sample
to 17.2 ng/µL with reduced EDTA TE buer in
a sterile, DNAse-free tube. For polyploid plants
and animals, dilute the sample to 23.0 ng/µL.
Prepare sucient material for the entire study. We
recommend one positive control per array plate.

2. Thaw samples and control.

Thaw the Genomic DNA Standard (Ref 103) and positive
control sample to temperature. To thaw, either:

• Place items on benchtop for one hour.

3. Quantify, then dilute gDNA.

a. Gently vortex (50% maximum), then centrifuge

samples.

b. Quantify each sample. The Quant-iT™ PicoGreen

dsDNA Assay Kit is recommended.

c. Using reduced EDTA TE buer, dilute each sample to a

concentration of:

• 11.5 ng/μL for human DNA samples

• 17.2 ng/μL for diploid plant and animal DNA
samples

• 23 ng/μL for polyploid plant and animal DNA
samples

d. Seal, vortex, then centrifuge.

4. Aliquot the diluted samples and the control.

Aliquot diluted samples and reference positive control DNA
to the deep well plate:

a. Aliquot 8.7 µL of each diluted gDNA sample (the

equivalent of 100–200 ng of gDNA depending on the
final sample type to be hybridized).

b. Aliquot 8.7 µL of the positive control sample. We

recommend including at least one positive control on
each plate.

c. Seal, then centrifuge.

5. Freeze or proceed. At this point you can:

• Store the sample plate at –20°C, or

• Proceed to DNA amplification for target preparation.

Note: If proceeding immediately to DNA amplification,
you can leave the gDNA sample plate at room
temperature.

6. Create a GeneTitan™ Array Plate Registration file.

Note: It is important to create and upload a GeneTitan

Array Plate Registration file with your sample information
before loading the array plate and hybridization tray in
the GeneTitan™ Instrument. Create (but not upload) this
file at the same time you prepare your plate of genomic
DNA. When your samples are ready for hybridization, scan
the array plate barcode and upload the file to GeneChip
Command Console™ (GCC).

GeneTitan™ Array Plate Registration file contains information
that is critical for:

• Data file generation during imaging.

• Tracking the experimental results for each sample that
is loaded onto an array plate.

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2 Axiom

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384HT and Mini 96 gDNA Sample Preparation Quick Reference