GenoExplorer™ Gene Expression
Biochips
Introduction to GenoExplorer™ Gene
Expression Products and Services
1. Facility and Environment of Fabrication and Laboratory
GenoExplorer™ Biochip (microarray) products and services
are manufactured and performed in a class 100 cleanroom facility
under stringent GMP and GLP protocols.
2. GenoExplorer™ Biochip Platform
GenoExplorer™ Biochips are established on a 25x75 mm high-quality
glass microslide with a 3D matrix. The DNA oligo probes are synthesized,
and then immobilized onto the chips. Only the full-length probes
attach to the chips, resulting in a high-quality performance indicated
by high reproducibility, signal to noise ratio, and sensitivity.
Scanners capture the fluorescent signals generated from the hybridization
assays.
3. Array Design
GenoExplorer™ chips contain the most comprehensive probe
content for gene profiling collected from all public domains. Chips
are manufactured based on the species, such as human, mouse, etc.
Probe designs have been computed and optimized based on their hybridization
thermodynamics. The chips offer the user the opportunity to perform
differential expression analyses between different samples. Multiple
positive and negative controls are included on the chips, such as
GAPDH, beta-actin, U6, and rRNA which also serve as loading controls.
Negative controls are used for chip background.
4. Isolation of Total RNA for Array Experiments
The main goal is to obtain good quality and sufficient amount of
RNA. Many laboratories use the Trizol reagent for isolation which
is recommended. However, each sample may result in different quality,
and further evaluation on those samples is necessary.
5. RNA Quality Evaluation for Gene Expression Experiments
High quality and sufficient concentration of RNA samples are crucial
for the experiments on microarrays. RNA quality can be evaluated
by visualizing the RNA on a gel, as well as by calculating the A260/A280
ratio. On a denaturing gel (or on an ordinary agarose gel in denaturing
buffer) the RNA should appear as two bright distinct bands that
represent the 28S and 18S ribosomal species. The 28S band should
be brighter than the 18S band. Tailing of these major bands down
the gel, or a background smear behind these bands that gets heavier
at lower molecular weights can indicate degradation of the RNA.
Degraded RNA will produce high background and low signal intensities
on microarrays. The presence of very sharp bands higher than the
28S ribosomal band can indicate the presence of excess DNA in the
sample, which can be removed by treatment with RNase-free DNase
I. The spectrophotometric ratio will also give an indication of
the purity of the RNA, and should be as close to 2.0 as possible.
Generally, ratios less than 1.7 indicate that the RNA may be contaminated
with other material and should be repurified, or perhaps run through
a column.
6. Amount and Concentration of RNA Samples
We use total RNA. The amount of RNA samples should be 1 µg/µl
or greater in DNase/RNase-free H2O. We require 1-2.5 µg of
total RNA for each testing sample(s).
7. Expression Profiling Technology
Total RNA in the amount of 5 µg per sample is required for
the assay. The labeled RNA is then used for the on-chip hybridization
assays under optimized conditions. The fluorescent signals from
hybridized targets are scanned and analyzed. The signal intensity
and background are computed. The analyzed data is stored and exported
to an application program for further analysis.
8. Submitting Your Samples
Before you submit your samples, please make sure 1) The RNA quality
has been properly evaluated. An agarose gel image of the RNA samples
is required. 2) Total RNA for each sample should be 1-2.5 µg
in the concentration of >0.5 µg/µl in DNase/RNase-free
H2O. 3) Samples should be sent on dry ice via an overnight courier
to the company’s address below. All RNA samples are stored
in –70°C upon arrival. 4) Complete the “Sample Testing
Submission Form” with the appropriate information.
Testing Samples Receivable
GenoSensor
4665 S. Ash Ave. Suite G-18
Tempe, Arizona 85282
USA
9. Final Report
We will send you the data in a compact disc that contains all scanned
chip images in tif image format, and raw data for each sample in
an Excel spreadsheet. The raw data is gridded and computed by standard
microarray analysis software, which is easily transferable to other
programs for further analysis. A data analysis report is also provided.
This data summary includes all gene names, fluorescent signal intensity
values of each probe, mean value of triplicate probes, standard
deviations, gene normalization, and fold changes between samples.
The chip layout that provides with the genes and their array positions
on the chips is also included on the CD.
10. Turnaround Time
Depending on number of chips and numbers required, the turnaround
time is typically one week to 10 days.
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