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Welcome to the Bio-Analytic Resource! Use the following links to access our data-mining tools, to search, or to contact us. These links are available on the menu bar at the top of each page for ease of navigation. The Main link will return you to this page.

Project Browser. Use this tool to access MIAME-compliant details (Brazma et al., 2001) for a particular project in the database. Projects are a collection of samples submitted by a researcher. In most cases projects come from researchers in the Department of Cell & Systems Biology at the University of Toronto.
Expression Browser. Use this program to perform electronic Northerns, that is, to see how up to 125 of your genes of interest are being expressed or are responding (how the genes are responding relative to the appropriate control) across all projects in our database, or in projects from the AtGenExpress Consortium.
Expression Angler. This tool will allow you to identify genes that exhibit similar expression or response profiles across all samples in the Bio-Analytic Resource data base, in 392 samples from NASCArrays, or in the AtGenExpress tissue expression data set. This program is useful for identifying potential interacting proteins or novel genes associated with a given process.
Contact Us. Any questions may be directed to the persons listed on this page.
Search. Search the database for a particular term.
Instructions. Although these tools are easy-to-use, some tips, background information, and other details are available here. Context-sensitive help is also available on the individual tool pages, too.

Background Information

The GeneChip facility in the Deptartment of Botany (now Department of Cell & Systems Biology) became operational in January 2003. It consists of a hybridisation oven, a fluidics station, a scanner and the necessary computer for controlling the latter two pieces of equipment. In addition we have two workstations for data analysis. Currently available for plant research is the ATH1 whole genome array which contains probe sets for querying the expression levels of approximately 22800 genes. Several other arrays are available commercially.

You may refer to http://www.affymetrix.com for an up-to-date list or the NetAffx tools at http://www.affymetrix.com/analysis/index.affx (free login required) may be used to check if a gene of interest is on the chip. Note too, that it is possible to have custom arrays designed. The costs are not insignificant, but perhaps several researchers could collaborate to have a chip designed with some of their genes of interest.

Procedure for Sample Preparation and Submission

  1. Individual researchers should plan their experiments themselves. Experimental design is important in order to be able to extract meaningful data from the results. Affymetrix recommends doing 3 biological replicates (i.e. 3 independent RNA isolations per time point, condition or mutant) . However, the reproducibility of the chips is very good (< 0.2% variability of independent hybs with the same RNA), so it might also be possible to pool the biological replicates and then do one hyb if you are just looking for candidate genes. It is also recommended to follow a protocol for RNA isolation. We recommend using RNAwiz for RNA preparation. With Trizol reagent (Gibco BRL, Cat#15596-026) yields and purity are lower, necessitating extra precipitation step(s). See the document on RNA reprecipitation using LiCl from Ambion at http://www.ambion.com/techlib/tb/tb_160.html. Use at least 0.5 M LiCl, keep at -20°C for more than 20 minutes, and spin at 4°C for more than 20 minutes. Yield tends to be low with the Plant RNAeasy from Qiagen, but the quality is good. Contact us for other extraction protocols for tissues that are rich in waxes or carbohydrates.

    For GeneChip analysis it is imperative that the RNA extracted be of high purity, displaying an OD260:OD280 ratio between 1.9-2.1. [Note: since the OD260:OD280 ratio is pH dependent, if the ratio appears to be unreasonably low (i.e. 1.5 or 1.6), an option is to spec it in 10 mM Tris-Cl, pH 7.5. The concentration; however, is measured most accurately in water. Sometimes it is useful to pH Milli-Q water to 7.5 and use this instead of Tris, if one can't spare another µl of RNA].

    To examine the quality and size distribution of the RNA sample, electrophorese 1 µg on a 1% agarose gel. High quality RNA samples will exhibit sharp rRNA bands, representing 28S and 18S RNA species. 23S and 16S rRNA bands may also be visible. Here is an example gel photo. The right lane contains the RNA (thanks to Wenzi Ckurshumova for the photo and above tip).

    For GeneChip experiments, a total of at least 10 µg of RNA is required. Not all this amount may be required, but we are erring on the side of caution.The concentration must be at least 0.5 µg per µl, so it is advised to resuspend in half the volume you think you would typically use then go from there

  2. Individual researchers purchase arrays through us (see below) and isolate total RNA themselves. Quality control parameters (OD260:OD280) should be provided on the Sample Submission Form along with the total RNA sample. Include a gel photo too. Please submit form in electronic format to Nicholas Provart, nicholas.provart@utoronto.ca along with a date when your samples will be ready. Note that the information will be required of you when you submit a microarray experiment for publication, according to the MIAME specifications, so you might as well do it now!

  3. Thanh will then perform the RNA labeling, prehyb, hybridisation, wash and scanning steps. The approximate cost for reagents for these steps is C per RNA sample, to be covered by the researcher.

  4. The individual researcher may then analyse the data from the experiment on his/her own, using Affymetrix's Data Mining Tool, publically available software, or GeneSpring. The key for the room where the data analysis workstations reside is available from Thanh.
    Material in the GeneChip Expression Analysis Technical Manual, Section 2: Eukaryotic Sample and Array Processing (Chapters 1, 3 and 4 therein) is relevant to data analysis. The manual may be downloaded at here or may be borrowed in hard copy from myself or Thanh. Material in the GeneChip Expression Analysis Data Analysis Fundamentals supersection is also relevant.

Cost Summary

Data Release

Diagram of Procedure