Products

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PyroMark Q96 ID System

Cat. No. / ID: 9001525

Instrument and software for Pyrosequencing analysis: includes installation, training, and 1-year warranty on parts and labor
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PyroMark Q24 System

Cat. No. / ID: 9001514

Instrument and software for Pyrosequencing analysis: includes installation, training, and 1-year warranty on parts and labor

Product Details

 

Resources

Supplementary Protocols (5)
Supplementary protocol using the EZ1 Advanced DNA Investigator Large-Scale Bone Card or the EZ1 Advanced XL DNA Investigator Large-Scale Bone Card
Supplementary protocol using the Flip Cap Rack and the EZ1 Advanced XL Flip Cap Card.
Protocol Files (4)
Search for QIAcube Standard Protocols
Purification of PCR products from 100-200 µl PCR samples

This protocol is for purification of double-stranded DNA fragments (100 bp to 10 kb) from PCR reactions of up to 200 µl sample volume (e.g., pooled samples) using the QIAquick PCR Purification Kit.

Sample Size 100-200 µl PCR samples
Elution volume 30-100 µl in increments of 10 µl, default 50µl
Applications Cleanup
Starting material Amplification reactions
Validation Reports (3)
For the EZ1 DNA Investigator Kit
  • BioRobot EZ1
  • EZ1 Advanced
  • EZ1 Advanced XL
Download Files (29)
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0008
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0013
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0004
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0002
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0009
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0011
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0010
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0005
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0003
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0012
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0001
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0006
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0007
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0014
Instrument methods file for use with PyroMark Q24 Software, PyroMark Gold Q24 Reagents, and PyroMark Q24 Cartridges marked with method code 0015
Brochures & Guides (7)
Fully automated low- to medium-throughput purification of nucleic acids
Introducing QIAseq
PDF (450KB)
Accelerate your NGS performance through Sample to Insight solutions
Kit Handbooks (4)
For automated purification of DNA from forensic and biosecurity samples using EZ1 instruments
EZ1 DNA Tissue Kit
For automated purification of DNA from tissue and other samples using EZ1 instruments
EZ1 Virus Mini Kit v2.0
For automated, simultaneous purification of viral DNA and RNA, and bacterial DNA, from 1-14 samples of serum, plasma, and CSF, urine, whole blood, and stool, transport media, respiratory samples, and dried swab samples using EZ1 instruments
EZ1 RNA Cell Mini Kit
For purification of total RNA from human and animal cells using EZ1instruments
EZ1 RNA Tissue Mini Kit
For purification of total RNA from easy-to-lyse human and animal tissues using EZ1 instruments
EZ1 RNA Universal Tissue Kit
For purification of total RNA from any type of human and animal tissue using EZ1 instruments
Instrument Technical Documents (1)
User-Developed Protocols (1)

This protocol has been adapted by the North Louisiana Criminalistics Laboratory from the pretreatment for epithelial cells mixed with sperm cells and is for the lysis and extraction of DNA from a sperm pellet after differential separation.

Software User Guides (1)

Publications

Genetic diagnostics of functional variants of the human dopamine D2 receptor gene.
Doehring A; Kirchhof A; Lötsch J;
Psychiatr Genet; 2009; 19 (5):259-68 2009 Oct PMID:19512960
Assessing combined methylation-sensitive high resolution melting and pyrosequencing for the analysis of heterogeneous DNA methylation.
Candiloro IL; Mikeska T; Dobrovic A;
Epigenetics; 2011; 6 (4):500-7 2011 Apr 1 PMID:21364322
Cell specific patterns of methylation in the human placenta.
Grigoriu A; Ferreira JC; Choufani S; Baczyk D; Kingdom J; Weksberg R;
Epigenetics; 2011; 6 (3):368-79 2011 Mar 1 PMID:21131778
CpG island hypermethylation of the neurofibromatosis type 2 (NF2) gene is rare in sporadic vestibular schwannomas.
Kullar PJ; Pearson DM; Malley DS; Collins VP; Ichimura K;
Neuropathol Appl Neurobiol; 2010; 36 (6):505-14 2010 Oct PMID:20831745
Systematic cross-validation of 454 sequencing and pyrosequencing for the exact quantification of DNA methylation patterns with single CpG resolution.
Potapova A; Albat C; Hasemeier B; Haeussler K; Lamprecht S; Suerbaum S; Kreipe H; Lehmann U;
BMC Biotechnol; 2011; 11 :6 2011 Jan 14 PMID:21235780

FAQ

What applications are offered on the QIAcube?

For an up-to-date list of available protocols, visit QIAcube Standard Protocols.

 

Click here for a virtual demonstration on how the QIAcube operates.

FAQ ID -1403
Where can I order the Streptavidin Sepharose beads for pyrosequencing?
The recommended Streptavidin Sepharose High Performance beads for pyrosequencing can be ordered at GE healthcare with the catalog no 17-5113-01.

The PyroMark Q48 Autoprep protocol uses magnetic streptavidin-coated Sepharose® beads (PyroMark Q48 Magnetic Beads), which bind to the biotinylated PCR strand.

PyroMark Q48 Magnetic Beads can be ordered at QIAGEN with the catalog no 974203.

FAQ ID -2850
Can I order the nucleotides from PyroMark Gold Reagents separately?
The nucleotides can only be ordered as part of the PyroMark Gold Reagents which also contain enzyme and substrate mix.
FAQ ID -2827
Where can I find the ordering information for QIAcube accessories such as Shaker Rack plugs, Rack labeling strips, Reagent bottle rack, 30mL Reagent bottles, Rotor Adapter holder, Rotor adapters, Sample tubes CB, Sample tubes RB and Spin column adapter rings?

Part numbers and prices for QIAcube accessories can be found by clicking here.

FAQ ID -3054
How many nucleotides of a homopolymer can be resolved in pyrosequencing?
In the range of 3-5 bases can be resolved depending on the sequence context and base. If it is possible sequencing of a homopolymer of more than 3-5 nucleotides should be avoided by resetting the sequencing primer.
FAQ ID -2871
I need to buy a new computer for my PyroMark Q96. Can I reinstall the PyroMark Q96 software on my new computer or do I need to purchase a new license?

If you need to purchase a new computer for your PyroMark Q96, or if you need to reinstall, upgrade, or replace the operating system (OS), you can reinstall the PyroMark Q96 software without purchasing a new license.

FAQ ID - 3472
How much space does the PyroMark Q24 instrument need?

PyroMark Q24 takes very little bench space, measuring only H420 x W390  x D525 mm.

 

FAQ ID -2098
Can the QIAcube heater/shaker be used independently from protocol runs?

Yes. The heated shaker on the QIAcube can be operated independently if the QIAcube is not performing a protocol run. For instructions on how to operate the shaker, refer to the QIAcube User Manual, chapter 5 'Operating Procedures, section 5.7 'Operating the shaker'.

The shaker can be used to heat and shake samples simultaneously, shake samples without heating, or heat samples without shaking.

 

See:  http://www.qiagen.com/products/catalogpagecontrols/header/qiacube/360/default.aspx for a virtual demonstration of the heater and shaker on the QIAcube.

FAQ ID -1422
What are the corresponding QIAGEN names for former Biotage instruments?

When looking at technical literature, pulse time settings, and other information, the following table can be used to determine the QIAGEN names for former Biotage instruments.

 

Biotage QIAGEN
PSQ 96 PyroMark Q96 ID
PSQ 96 MA PyroMark Q96 ID
PSQ HS 96 PyroMark Q96 MD
PSQ HS 96A PyroMark Q96 MD Automated
FAQ ID -2285
What is a PyroMark instrument method or instrument code?

An instrument method or instrument code encodes the individual pulse time settings of specific cartridge lot batch. These pulse time settings change when e.g. a new batch of capillaries is used with slight variations in the needle diameter. For larger diameters, the pulse settings are lowered to dispense the correct volume of liquid. In addition, the viscosity of enzyme and substrate mixes can change which influences dispensing volumes.

The individual instrument method/code number is printed on the cartridge label. The corresponding methods/code settings can be downloaded as a file from the respective instrument webpage and opened in the PyroMark application software.

FAQ ID -2941
What is the reason for a high substrate peak in the pyrosequencing pyrogram?
Usually pyrophosphate or dATP/ATP contamination in the sample or in the buffer can cause a high substrate peak. Large amounts of pyrophosphate are generated in the PCR reaction and might be carried over to the sequencing reaction. Check the PyroMark buffers and reagents and use new ones.
FAQ ID -2879
How can I load new protocols onto the QIAcube?

Visit the QIAcube Protocols page to search for protocols. The instructions for transferring a protocol to the QIAcube are listed below.

To transfer new protocols to the QIAcube, first save them to the supplied USB stick.

  1. Select the protocol to be downloaded and save it to the “New_Protocols” folder on the USB stick.
  2. Make sure that the QIAcube is switched on.
  3. To install the protocol, connect the USB stick to the QIAcube via the USB port, which is located at the front right of the instrument.
  4. In the main menu, press “Tools”.
  5. Select “Data Exchange” by pressing the "Up Symbol" or the "Down Symbol" to scroll through the list until it is highlighted and then press “Select”.
  6. To transfer protocols to the QIAcube, select “Protocols” and “Load from USB”.
  7. Start the download procedure by pressing “OK”. A message will confirm that the protocols have been successfully transferred to the QIAcube.

Note: Protocols already installed on the QIAcube that have the same name as downloaded protocols will be overwritten.

    8. When the protocols have been successfully transferred to the QIAcube, press “OK”.

 

 

 

FAQ ID -1421
How does the built-in QC control for complete bisulfite conversion of DNA work on PyroMark Q24?

Data generated with the methylation analysis software (detection and quantification of CpG sites) on PyroMark Q24 contains unique features that act as quality control for complete bisulfite conversion of DNA. When the assay encounters a C not followed by a G (C unmethylated), that C should be fully converted to T if the bisulfite treatment upfront was successful. Subsequently, it should be presented in the pyrogram as T=100%. This acts as a useful quality control for full conversion of unmethylated C residues during bisulfite treatment and PCR.

FAQ ID -2095
Are run reports and/or log files available on the QIAcube?

Yes. At the end of each protocol run on the QIAcube, a report file is generated that contains important information such as number of samples, liquid volumes measured during the load check, and run time. A log file is also generated that contains information about instrument actions, such as movement of the robotic arm.

 

FAQ ID -1412
What are the features of the new PyroMark Q96 ID software 2.5?

This software will replace the old PyroMark Q96 ID software version 1.0. The PyroMark Q96 ID software version 2.5 contains four assay setup and analysis modes which are AQ, CpG, SNP and SQA. It is based on the Pyromark Q24 application software and is a file based software (runs are not stored in an ORACLE database). It replaces the PyroMark CpG software since a CpG mode is included. There is no SNP multiplexing support and no alignment functionality (as found in software version 1.0).

FAQ ID -2867
Which end of the PCR primer for pyrosequencing should be biotinylated?
In Pyrosequencing, the 5' end should be biotinylated, regardless of whether the forward or reverse primer is biotinylated. You can order pyrosequencing primers here.
FAQ ID -2839
Can the QIAcube centrifuge be used independently from protocol runs?

Yes. The QIAcube centrifuge can be operated independently if the QIAcube is not performing a protocol run. For instructions on how to operate the centrifuge, refer to the QIAcube User Manual, chapter 5 'Operating Procedures, section 5.8 'Operating the Centrifuge'.

 

 

FAQ ID -1423
What kind of shaker should be used for the pyrosequencing binding step?
Shaking conditions are 1400 rpm at room temperature. Optimal results are obtained with 2mm orbital diameter.
FAQ ID -2837
Which analyses can be performed with the PyroMark Q96 ID software version 1.0?
The PyroMark Q96 ID software version 1.0 contains a SNP mode for simplex and multiplex entries (and Allele Quantification analyses can be performed for simplex entries) and an SQA mode for de novo sequencing analysis. This software version does not contain a CpG mode for methylation analysis, however the PyroMark CpG software v1.0 can be used.
FAQ ID -2845
Is it necessary to place the lids of the elution tube and column into the slots of the rotor adapter during processing on the QIAcube?

Yes, this is absolutely necessary. If the lids are not securely fixed in the rotor adapter slots during processing on the QIAcube, they will become detached from the elution tubes and columns during centrifugation and will collect at the bottom of the centrifuge chamber.

 

FAQ ID -1415
What should be the single peak height for the PyroMark Control Oligo on the different PyroMark instruments?


PyroMark Q24: The mean single peak height is 95 +/- 55 RLU
PyroMark Q48: The mean single peak height is 70 +/- 40 RLU
PyroMark Q96 ID: The mean single peak height is 35 +/- 10 RLU
Pyromark Q96 MD: The mean single peak height should be at least 350 RLU

FAQ ID -2852
Are there any specific cleaning solutions recommended for the QIAcube?

The following disinfectants and detergents are recommended for cleaning the QIAcube:

  • Mikrozid® Liquid (Schülke & Mayr GmbH) — ethanol-based disinfectant for spraying onto surfaces (consists of 25 g ethanol and 35 g 1-propanol per 100 g Mikrozid Liquid)
  • RBS®-35 Detergent Concentrate (Pierce Chemical Co.) — anionic and nonionic surfactant based detergent for cleaning the touch screen and worktable door
  • RNaseZap® (Ambion) — for complete removal of RNase contamination from glass and plastic surfaces
  • 0.1 M NaOH — for cleaning the worktable and racks to remove RNase contamination (alternative to RNaseZap)
  • Note: Alcohol is not recommended for cleaning of the door.

Europe 

Lysetol® AF (Gigsept® Instru AF) (Schülke & Mayr GmbH) — disinfectant for submerging worktable items (consists of 14 g cocospropylene-diamine-guanidine diacetate, 35 g phenoxypropanols, and 2.5 g benzalkonium chloride per 100 g Lysetol AF, with anticorrosion components, fragrance, and 15–30% nonionic surfactants).

 

USA

DECON-QUAT® 100 (Veltek Associates, Inc.) — quaternary ammonium salt based disinfectant concentrate for submerging worktable items (contains 5% alkyldimethylbenzylammonium chloride and 5% alkyldimethylethylbenzylammonium chloride).

Note: If you want to use disinfectants different from those recommended, ensure that their compositions are similar to those described above.

FAQ ID -1416
What is the use of the PyroMark Q24 Validation Oligo?
The PyroMark Q24 Validation Oligo was developed to check the performance of the PyroMark Q24 system. It consists of two biotinylated oligonucleotides that only differ in one position (A or G) of the sequence. By mixing different proportions of the two oligonucleotides in replicates the linearity, bias and reproducibility of the system can be determined.
FAQ ID -2855
What is the maximum number of samples for the QIAcube variant of the PAXgene Blood miRNA protocol for a single run?

Up to 12 samples can be processed in a single run.

FAQ ID - 3482
How can I get software updates for the QIAcube?

New software or software updates can be accessed via the QIAcube Web Portal at www.qiagen.com/MyQIAcube. To access them directly, go to User Support.

 

 

FAQ ID -1413
Can purification columns from other suppliers be processed on the QIAcube?

No. The QIAcube is designed for fully automated processing of up to 12 samples using QIAGEN spin-column kits. Do not use spin columns manufactured by other suppliers with the QIAcube. Damage caused by use of other types of spin columns will void the instrument warranty.

 

See:  http://www.qiagen.com/products/catalogpagecontrols/header/qiacube/360/default.aspx for a demonstration on how the QIAcube is used with the RNeasy Mini Kit for automated purification of RNA.

 

FAQ ID -1406
Can I program my own protocols for the QIAcube?

No. Modified QIAGEN protocols or custom protocols developed to meet your specific requirements can be requested and purchased from QIAGEN.

1. Go to www.qiagen.com/MyQIAcube and enter the required information. If our Application Specialists require additional information, they will contact you directly.

2. You will be informed by e-mail when your customized protocol is ready for download.

3. Click the link in the e-mail to go to the download page.

FAQ ID -1405
What is the sensitivity limitation for pyrosequencing?
In general, the standard claim for pyrosequencing sensitivity is about 5% which is also published in many papers. The actual sensitivity limit is assay dependent and has to be determined individually.
FAQ ID -2840
When do I have to change the pulse settings/methods in a pyrosequencing run setup?
Always check for the actual method/code number printed on the cartridge label. Make sure that you choose this method/code number when setting up the pyrorun in the application software. If this method cannot be selected automatically in the application software, you can download the method/code file from the instrument webpage.
FAQ ID -2942
Which PyroMark Gold Q96 Reagent should be used for which instrument and application?

PyroMark Gold Q96 Reagents:

  • PyroMark Gold Q96 SQA Reagents (1 x 96) should be used for performing SQA analyses on the PyroMark Q96 ID. It can also be used to supplement the PyroMark Gold Q96 Reagents (5 x 96) when long runs lead to a shortage of nucleotides.
  • PyroMark Gold Q96 Reagents (5 x 96) should be used for SNP and CpG analyses on the PyroMark Q96 ID and MD instruments. On the PyroMark Q96 MD, the kit contains enough for 15x96 plates.
  • PyroMark Gold Q96 CDT Reagents (6 x 96) should be used only with CDTs on the PyroMark Q96 MD instrument, for performing SNP and CpG anslyses. The nucleotides are pre-diluted for use with CDTs, so this is not to be used on ID instrument or with NDTs on the MD.
  • PyroMark Gold Q96 Reagents (50 x 96) should be used only with the PyroMark Q96 MDA (automated option) for SNP anslyses. Not for use with the PyroMark Q96 ID or non-automated MD.

See the PyroMark Gold Q96 Reagents Handbook for additional information.

FAQ ID -2836
How do I reduce background peaks in the pyrosequencing pyrogram?
There are several reasons for a high assay background; the template can form secondary structures which are extended or the primers itself form dimmers which serve as template. Perform accurate sequencing controls (e.g. PCR or sequencing primer only) as recommended in the PyroMark User Manual to observe this kind of background. In addition, an unspecific priming of primer to template or unspecific annealing of sequencing primer to template might also be a background cause. Please check your complete primer design and if needed, perform a redesign. Try to lower the primer concentration as possible to avoid excess of primer.
FAQ ID -2877
What is the sample throughput of Pyrosequencing systems?

PyroMark instruments offer a range of throughput scales. The PyroMark Q24 can process 1–24 samples in parallel, the PyroMark Q48 Autoprep 1-48, the PyroMark Q96 ID 1–96, and the PyroMark Q96 MD 1-96, or the automation option enables automated processing of ten 96-well plates. The sample processing speed depends on the number of nucleotide dispensations necessary for the programmed analysis. Twenty dispensations take approximately 24 minutes on all instruments; thus, 96 samples are typically processed in 10-100 minutes.

 

 

FAQ ID -2215
What is the concentration of PyroMark Control Oligo?
PyroMark Control Oligo has a concentration of 20µM and is delivered in a volume of 50µl. Two tubes of 10x dilution buffer (2x 1.7ml) are delivered with the control oligo.
FAQ ID -2846
What is the pyrosequencing data exchange tool for?
This tool is part of the old PyroMark Q96 ID/MD systems that run with the ORACLE database. The tool enables exporting and importing run files from the ORACLE database.
FAQ ID -2864
Can the QIAcube be connected to a laboratory information management system?
No, it is not possible to connect the QIAcube with laboratory computer networks.
FAQ ID -1411
How many times can vacuum troughs be re-used with the PyroMark Vacuum Preparation Stations?
There is no precise recommendation how many times these troughs on the PyroMark Vacuum Preparation Stations (Q24 and Q96) can be re-used. It depends on the individual handling and cleaning (with water).
FAQ ID -2848
Which heating block is recommended for the pyrosequencing annealing step?
A heating block that can heat up to 80-90 °C is recommended. A solid block is preferable. For the PyroMark Q24 the surface area must be 50mm x 60 mm and for Pyromark Q96 ID/MD 120mm x 80 mm.
FAQ ID -2838
Can I reinstall the PyroMark Q24 software on my new computer or following an operating system upgrade, or do I need to purchase a new license?

If you need to install the PyroMark Q24 software on a new computer replacing your old one, or after you reinstall or upgrade the operating system, you can reinstall the PyroMark Q24 software without purchasing a new license.

FAQ ID - 3473

What is the PyroMark Q96 Data Converter?
PyroMark Q96 Data Converter 1.0 is a tool for exporting pyrosequencing runs from PyroMark Q96 ID ORACLE database. It converts the runs into a file format that is compatible with the new PyroMark Q96 ID software version 2.5.
FAQ ID -2868
What concentration should be used for the sequencing primer in pyrosequencing?

Usually the sequencing primer is used at 0.3µM in annealing buffer but some assays might require additional optimization of the sequencing primer concentration.

 

For PyroMark Q24 and PyroMark Q96 MD the final concentration of the sequencing primer is 0.3µM and for PyroMark Q96 ID 0.4µM.


The PyroMark Q48 Autoprep dispenses the sequencing primers for annealing. The final concentration of sequencing primers in a well is 0.8µM, but may be adapted to optimize assays.

 

FAQ ID -2826
What dedicated QIAcube Kits are available?
What are the dimensions and weight of the QIAcube?

The QIAcube has the following weight and dimensions:  

  • Weight: 70.5 kg (155.4 lb.)
  • Width: 65 cm (25.6 in.)
  • Depth: 62 cm (24.4 in.)
  • Height (door closed): 57 cm (22.4 in.)
  • Height (door open): 81 cm (32 in.)
FAQ ID -1407
Where to find explanations to the warning given by the PyroMark software after run data analysis?
The PyroMark Q24 application software contains under Help (Press F1) a Pyromark Q24 Software user guide with all essential information about warnings and software features. The Pyromark ID/MD software also contains a softwareware guide under help. Moreover, the individual instrument user manuals contain helpful information in the troubleshooting section.
FAQ ID -2874
Which purity grade is recommended for pyrosequencing primers?
Only the biotinylated primer needs to be HPLC purified whereas the other primers require standard desalting only.  Pyrosequencing primers can be ordered here.
FAQ ID -2832
What is the reason for split peaks appearing in between dispensations on my pyrosequencing pyrogram?
The PyroMark cartridge needle can be blocked or damaged. Clean the cartridge or exchange with a new one. Check for correct reagent cartridge and cartridge method used in the run. Check if the reagent cartridge cover was closed properly. Make sure that the cartridge was dry after cleaning because nucleotide droplets might be caught at the needle tip and fall down at any time. or exchanged.
FAQ ID -2881
How can I decontaminate the QIAcube system?

Please refer to section 6.5 'Decontaminating the QIAcube' in chapter 6 'Maintenance Procedures' of the QIAcube User Manual.

 

 

 

 

FAQ ID -1419
Will dUTP in a PCR reaction affect pyrosequencing?
In general, dUTP/UNG treatment should work for pyrosequencing in order to reduce contamination risk with PCR amplicons from previous PCR’s.
FAQ ID -2843
Is the CpG software included in the PyroMark instruments to study methylation status?
The PyroMark Q24 software and the new PyroMark Q96 ID software version 2.5 support CpG analysis in the CpG mode. The ORACLE-based PyroMark Q96 ID software version and PyroMark Q96 MD software do not support CpG analysis. In this case an independent, additional software is needed which is the PyroMark CpG software version 1.0.
FAQ ID -2842
How long does a single run on the PyroMark Q24 take?

PyroMark Q24 analyses up to 24 individual samples in a single run. The time needed to perform a run depends on the number of nucleotides dispensed in the specific assay. A typical SNP assay contains 15 dispensations and a typical CpG assay 45. Since the estimated dispensation time is 1 minute/nucleotide, a typical SNP run takes 15 minutes to perform and a typical CpG assay takes 45 minutes.

FAQ ID -2096
Can the QIAcube rotor and/or buckets be removed for cleaning?

Yes, rotor and/or buckets can be removed from the QIAcube for cleaning. For information on how to remove these parts, refer to the QIAcube User Manual.

 

FAQ ID -1410
Can QIAcube protocol runs be interrupted, and subsequently be continued to completion?

You can stop a QIAcube protocol if there is an emergency by pressing “Cancel”. To confirm that you want to stop the protocol run, press “OK”. To cancel the 'stop protocol' command, press “Cancel”.

Note: If a protocol run is stopped, the run cannot be restarted. The samples must be processed manually. The last protocol step performed by the QIAcube will be displayed in the touch screen. For more information, refer to the QIAcube User Manual.

FAQ ID -1414
Do I need to buy special kits for the QIAcube?

The corresponding manual kits can be used for all available protocols on the QIAcube. The manual kits contain sufficient buffers and reagents to process the number of spin columns provided with the kit.

In addition to the purification kit, Rotor Adapters (10 x 24), cat. no 990394, and the required disposable tips:

  • Filter-Tips, 200 µl (1024), cat no. 990332,
  • Filter-Tips, 1000 µl (1024), cat no. 990352,
  • Filter-Tips, 1000 µl wide-bore (1024), cat no. 990452,

need to be ordered separately. To find out which filter-tips are required for a specific application, refer to the corresponding protocol sheet.

FAQ ID -1402
Can the QIAcube rotor adapters be autoclaved?
No. Autoclaving the rotor adapters will cause the plastic to deform and the rotor adapters will no longer fit properly into the centrifuge buckets.
FAQ ID -1409
What is the reason for signals ceasing in the middle of a pyrosequencing run?
The cartridge needle can be blocked or damaged causing a dispensation error. Clean the cartridge following the guidelines or repeat the run with a new cartridge. On the other hand if high amounts of template have been used resulting in very high signals (>100 RLU), the substrate for the sequencing reaction might be depleted. In this case template conditions should be optimized.
FAQ ID -2875
3323 - Do you have a protocol for the AllPrep DNA/RNA/Protein Mini Kit on the QIAcube?
A custom protocol can be requested through the online QIAcube protocol request page: <https://www.qiagen.com/qiacube/customized/protocoladd.aspx >.
FAQ ID - 3323

 

What kind of reading length can I expect when using Pyrosequencing technology for sequence analysis?

Typical reading length using Pyrosequencing technology is 40-60 bases. However, as with any sequencing technology, the maximum read length will depend on template secondary structure, base content, quality of PCR-product, and other parameters.

Depending on the sequence to be analyzed, highly accurate read lengths of 140 or more bases can be obtained in just a single reaction with the Q48 PyroMark Autoprep.

 

 

FAQ ID -2216
What is the recommended amplicon size for CpG assays?
The amplicon length should be short (<200bp). This is critical especially for DNA from FFPE tissue which is often degraded by the fixation so that short fragments are easier to amplify. Moreover, the DNA suffers from harsh bisulfite treatment and might receive further double strand breaks. Therefore the amplicon size should be kept as short as possible.
FAQ ID -2825
Can unused wells in a pyrosequencing plate be used in the next run?
In principle it’s possible to use so far unused pyrosequencing wells for the next run and leave the already used wells empty. However, due to contamination risk when cleaning and handling plates QIAGEN does not recommend this.
FAQ ID -2872
Do I need to discard partially used QIAcube tip racks?

No. The QIAcube is equipped with an optical sensor, which determines the tip size and the number of tips loaded onto the worktable prior to starting a run.

The QIAcube can use partially used tip racks from previous runs, provided that the racks contain sufficient tips to process the number of samples loaded for the current run.

FAQ ID -1418
Do the samples require pretreatment before starting the manual protocol or before loading onto the QIAcube instrument?
Yes. Samples have to be incubated at least 2 h at room temperature (18-22°C). Incubation of the PAXgene Blood RNA Tube overnight may increase yields slightly in some cases. Room temperature storage for up to three days has no effect on the RNA profile. During the preparation of the RNA the manual pretreatment consist of a centrifugation and a pellet resuspenion step (see the PAXgene Blood miRNA Handbook ).
FAQ ID - 3479
How accurate and reliable is PyroMark Q24 in mutation analysis?

PyroMark Q24 uses Pyrosequencing technology for mutation analysis and provides a built-in quality control in each run. By sequencing nucleotides flanking the mutation of interest, the researcher gets confirmation that the assay was made at the correct position. In addition, blank dispensations are included in the assay providing a negative control of the run. 

FAQ ID -2094
How do I prevent a drifting baseline in my pyrosequencing pyrogram?
Let the PyroMark instrument warm up (about 60 minutes) to adapt to room temperature before use. Make sure the ambient room temperature is within range 18-28°C.
FAQ ID -2878