Microarray

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RNA Quality Assurance Instructions

Before submitting RNA samples for labeling and hybridization, please follow the procedures below.

RNA must be prepared and examined using the following assays.

1. Isolate total RNA using Trizol (GIBCO,BRL) and clean using RNeasy Kit (Qiagen). When using mRNA it must be isolated from total RNA using Oligotex columns (Qiagen) or oligo DT cellulose columns (GIBCO,BRL).
   
   
2. Measure the optical density (OD) of total RNA and mRNA samples including OD₂₆₀, OD₂₈₀, OD₂₆₀ /OD₂₈₀, and calculate the concentration (µg/µl) of your samples. Submit your samples with all the values above and the dilution factor that you used to take these readings.
   
   For each mRNA sample, please send at least 5 µg at a concentration of 0.25 µg/µl.
   
   For total RNA samples
   For Eppendorf, Agilent or Numblegen arrays send 12 µg at a concentration of 1 µg/µl
   For Affymetrix arrays where standard one-cycle cRNA synthesis is performed, send at least 5 µg at a concentration 0.25 µg/µl.  For smaller amounts of total RNA, a two-cycle cRNA synthesis is performed.   At least 0.25 µg is necessary when choosing this option.
   
   Users outside of UAlbany East Campus must send RNA samples in ethanol on dry ice or as a dry pellet.
   
   
3. Visualize each total RNA sample by running on an agarose gel containing 1.1% formaldehyde and photograph. Please attach a photo of the results (please remember to clearly label each lane with the sample label).
   
   Please label each tube of RNA sample prepared in DEPC-H2O carefully and complete all the details on the RNA submission form with the volume and concentration. This form must be faxed to 518-591-7211 or hand delivered to Center for Functional Genomics, 1 Discovery Dr., Room 328, Rensselaer, NY 12144.

Our protocol for RNA Quality Control

To obtain high quality data from your gene chip experiment, it is essential to start with high quality total RNA. If RNA is degraded or otherwise unsuitable for microarray application, investigators will be notified immediately.

Spectrophotometric Analysis:
A small amount of your sample will be assayed with a Nanodrop ND-1000. We require a concentration of at least 1 mg/mL. The OD 260/280 is calculated to estimate the purity of the RNA. A ratio close to 2.00 indicates a high percentage of ribonucleotide.

Electrophoretic Analysis:
To assess the integrity of your total RNA, we will test it with the Agilent Technologies 2100 Bioanalyzer Lab-on-a-chip system. This assay is similar to gel electrophoresis in concept, but it is cleaner, more efficient, and only requires a very small amount of sample.

Agilent's Lab-on-a-chip technology

A small amount of sample is loaded into the wells in the chip and electrodes cause the RNA to move through microchannels filled with a sieving polymer and fluorescent dye. Fluorescence signal is plotted against run-time to generate an electropherogram or translated to gel-like images. Quality metrics such as 28S/18S ratio and RIN (RNA integrity number) are considered before the sample proceeds through the queue.

Instrumentation | Fees for Services | Sample Preparation, Handling and QC | Procedure for Submitting Samples
Turn around Time | Frequently Asked Questions | Useful Links