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Roche NimbleGen’s proprietary array manufacturing technology allows fast and cost-effective synthesis of custom-designed arrays. Custom-designed arrays may be ordered for delivery (for hybridization in your own lab) for any organism.
Custom designs may also be ordered for full service analysis if they meet the design principles described below. These principles will ensure best performance using the standard NimbleGen array processing protocols.
Custom Design Principles for Full-Service Workflow
- Transcriptome GC content is between 35 and 60%
- Probe selection is ORF-based with 3’-bias
- Probe length is 60-mer for eukaryotes and 45-60-mer for prokaryotes
- Transcriptome size allows majority of transcript to have ≥ 3 probes per target
Please contact Roche NimbleGen Sales for a quotation or contact NimbleGen Technical Support with any questions about gene expression custom arrays.
Ordering Information 
Literature
For a complete listing of literature covering all Roche NimbleGen products and services please visit our literature page.
FAQ
| Hide All Topics Show All Topics |
| Multiplex |
| What are the advantages of the Roche NimbleGen multiplex array formats? |
The NimbleGen 12x135K and 4x72K array formats allow simultaneous hybridization of 12 or 4 samples, respectively, on a single slide. You can conveniently run either replicates of the same sample or multiple samples on the same slide, thus improving interarray reproducibility and reducing cost. |
| How many features are on each array of a Roche NimbleGen multiplex slide? |
Each array on the NimbleGen 4x72K slide has 72,000 features and each array on the 12x135K slide has 135,000 features. |
| Do you offer multiple probes per transcript in the NimbleGen multiplex formats? |
Yes, each transcript is interrogated by 3 or more probes, depending on the complexity of the organism. Please see our catalog for more details for each organism. |
| What kind of controls do you include on each array to ensure accurate sample loading and to confirm the integrity of the experiment? |
Each array contains probes for Sample Tracking Controls (STCs) that are used for the following purposes:
a) To identify the samples after hybridization; each labeled cDNA sample is spiked with a unique Sample Tracking control before loading.
b)To ensure that there is no sample mix up between arrays. |
| Do all arrays on a Roche NimbleGen multiplex slide have the same design? |
Yes. Currently all the arrays on a gene expression multiplex slide have identical designs. |
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| Array Design |
| Do you include microRNAs or non-coding RNA in your catalog designs for expression? |
Currently, no microRNA or non-coding RNA regions are used for NimbleGen gene expression array designs. |
| What is the probe length in gene expression arrays? |
NimbleGen expression arrays contain 60-mer probes. |
| Do you offer catalog and custom array designs? |
Yes, we offer both catalog and custom array designs for differential expression analysis. Please check our website for the current catalog. If you do not see your organism, please contact the Account Manager for your region to discuss a custom design. |
| Are your gene expression probes designed with a 3' bias? |
Yes, probes are designed with some 3' bias. Our expression arrays are designed by first determining the transcript terminator as annotated in NCBI. We then take 1500bp upstream of that location and design our probes within that region. |
| Which strand is represented on the array? |
The sense strand is represented on our arrays. |
| Can Roche NimbleGen design a custom expression tiled array for my specific research needs? |
Yes, Roche NimbleGen can design and deliver to you custom tiled expression microarrays with probe spacing and probe length that best fits your experimental goals. Please contact your local sales representative to discuss a custom design. |
| Are Roche NimbleGen designs up-to-date with the latest genome builds? |
Yes, Roche NimbleGen technology allows the rapid production of new array designs based on the latest genome builds. |
| Are designs based on earlier genome builds available? |
Yes, please contact the Account Manager for your region. |
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| Sample Preparation |
| What is the recommended protocol for isolating RNA? |
We recommend the RNeasy Mini Kit (Qiagen, Cat. No. 74104) or the TRIzol Reagent (Invitrogen, Cat. No. 15596-026)). If using TRIzol-isolated RNA for cDNA synthesis, it is beneficial to perform a secondary cleanup step using RNeasy Mini Kit. Immediately after the ethanol precipitation step in the TRIzol procedure, proceed with the RNeasy Mini Kit according to the manufacturers recommendations. |
| Can I use a different RNA isolation method than recommended? |
Any RNA isolation method that provides RNA which meets the purity and integrity criteria outlined by NimbleGen can be used. |
| How do you assess the quality of RNA and cDNA samples? |
Pure and intact RNA and cDNA should have A260/A280 and A260/A230 ratios of at least 1.8. Additionally, they should appear intact when analyzed by gel electrophoresis or using a Bioanalyzer (Agilent). |
| How much RNA or cDNA do you need for gene expression analysis? |
This depends on whether you are sending the samples to Roche NimbleGen for full service or if you are doing the experiment in your lab, and whether your sample is from eukaryotes or prokaryotes. When performing experiments in your lab using the Roche NimbleGen-recommended protocol, 10 μg total RNA, 1 μg mRNA, or 1 μg cDNA is required. For service, the amounts required are listed in the table below.
| |
Total RNA* |
mRNA* |
cDNA |
| Eukaryote |
20μg |
5μg |
2.5μg |
| Class I Prokaryote |
20μg |
- |
2.5μg |
| Class II and III Prokaryotes |
30μg |
- |
5μg |
| *Please note that the Roche NimbleGen procedure for sample processing does not include RNA amplification. |
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| What is the expected cDNA yield from 10μg total eukaryotic RNA? |
Using our standard protocol, we routinely get at least 5μg cDNA from 10μg total RNA. |
| If I do not have 10μg of RNA, what method for RNA amplification can I use, and can I submit the cDNA samples for full service? |
cDNA synthesis methods that have been validated to perform well with NimbleGen eukaryotic gene expression service are listed below, together with the starting total RNA amount. Roche NimbleGen will also accept cDNA from amplified eukaryotic RNA. Please see the recommended RNA amplification kits below.
- TransPlex® Whole Transcriptome Amplification Kit (Sigma, cat. no. WTA1) with 50 ng total RNA
- TransPlex® Complete Whole Transcriptome Amplification Kit (Sigma, cat. no. WTA2) with 50 ng total RNA
- RiboMultiplier™ Sense-RNA Amplification Kit (Epicentre, cat. no. RM80510) followed with dsDNA Conversion Kit for RiboMultiplier™ sRNA (Epicentre, cat. no. RMD80625) with 100 ng total RNA
- Ovation® RNA Amplification System V2 (NuGEN, cat. no. 3100) with 20 ng total RNA
- WT-Ovation™ Pico System (NuGEN, cat. no. 3300) with 20 ng total RNA
These cDNA preparation methods have NOT been validated for prokaryotic samples. |
| What kind of samples are recommended for hybridizing on NimbleGen gene expression arrays? |
We recommend using Cy3-labeled cDNA sample on our gene expression arrays. Please contact technical support for your region to obtain a recommended protocol for preparing Cy3-labeled cDNA. |
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| Data Analysis |
| What method of normalization does NimbleGen use for gene expression? |
We normalize expression data using quantile normalization. See Bolstad B, et al. (Bioinformatics, 19:185-193, 2003), A comparison of normalization methods for high density oligonucleotide array data based on bias and variance. |
| How does NimbleGen generate gene calls from gene expression arrays? |
Gene calls are generated using the Robust Multichip Average (RMA) algorithm. See Irizarry R. et al. (Biostatistics 4:249-264, 2003), Exploration, normalization, and summaries of high density oligonucleotide array probe level data. |
| What software do you recommend for further analysis of gene expression data? |
NimbleGen provides gene expression values in calls file that can be conveniently imported into multiple gene expression analysis software such as ArrayStar (DNAStar). Currently, with the purchase of a full service project, you will obtain a 30-day demo version of ArrayStar along with your data. Array delivery customers can also request this 30-day demo version from NimbleGen. |
| Can NimbleGen gene expression data be used with publicly available software packages? |
Several analysis packages that have been used with NimbleGen gene expression data are:
1. NMPP: Described in Bioinformatics (2006); 22: 2955 - 2957
2. MeV: Described in J. Exp. Biol. (2007); 210: 1507 - 1517.
3. R: Available at www.bioconductor.org, requires data in XYS format. |
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| Array Service |
| How much RNA or cDNA do you need for gene expression analysis? |
| |
Total RNA* |
mRNA* |
cDNA |
| Eukaryote |
20μg |
5μg |
2.5μg |
| Class I Prokaryote |
20μg |
- |
2.5μg |
| Class II and III Prokaryotes |
30μg |
- |
5μg |
| *Please note that the NimbleGen procedure for sample processing does not include RNA amplification. |
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| When I use NimbleGen full service option, what deliverables will I receive ? |
NimbleGen delivers the following data files and software to full service customer:
- Microarray Design file containing probe sequences (ndf)
- Gene Description file (ngd)
- Raw Data files (calls and pair)
- Normalized Data files (RMA_calls and RMA_pair) |
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| Array Delivery |
| Can I purchase NimbleGen arrays for use in my own lab or core facility? |
Yes, all catalog designs currently listed on NimbleGen website and any custom design are available for delivery. |
| What kits are available for use with NimbleGen microarrays? |
NimbleGen offers One-Color DNA Labeling Kits, Hybridization kits, and Wash Kits to facilitate efficient and specific hybridization to our expression arrays. See our Kits & Consumables web page for more details about these kits. |
| What resolution scanner do I need? |
The NimbleGen MS 200 scanner set at 5 micron resolution is optimal for 385K arrays. The NimbleGen MS 200 scanner set at 2 micron resolution is optimal for 2.1M arrays. |
| Can I scan NimbleGen arrays using my Agilent scanner? |
The 5 micron Agilent scanner can work with NimbleGen arrays. |
| What software do I need to extract my data from a scanned image? |
Roche NimbleGen’s NimbleScan software can be used to perform feature extraction from scanned images. |
| Does NimbleGen offer training for array delivery customers? |
Yes, Roche NimbleGen Workshops are intensive, 2 to 4-day training programs designed to provide instruction in both the theory and practice of genomic and epigenomic research techniques using Roche NimbleGen high-density DNA microarrays. Click here to find out more. |
| When I order microarrays from NimbleGen, what deliverables will I receive? |
For microarray orders, NimbleGen delivers the following items to the customer:
- NimbleGen arrays
- User’s Guide
- Microarray Design file containing probe sequences (ndf)
- Gene Description file (ngd) |
| How much sample do I need to run one microarray analysis in my lab? |
When using NimbleGen-recommended protocol for cDNA synthesis and labeling, 10 μg total RNA, 1 μg mRNA, or 1 μg cDNA is required for each array. |
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