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  Genomic Services
• Oligo Synthesis
• Gene Expression
• Real-Time PCR
• Pyrosequencing
• Next Gen Sequencing
• DNA Sequencing
   & Fragment Analysis
• Single Cell Genomics
• RNA/DNA QC
‡ Bioanalyzer QC
‡ Fragment Analyzer QC
  Protein Services
• Peptide Synthesis
• Mass Spectrometry
• Protein Analytics
‡ Mass Mapping
‡ Edman Sequencing
• Biacore
• Antibody
 
 
Located in rooms
B065 and B017
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Gene Expression
 

DATA ANALYSIS

Scanning and/or Full Service: (back to top)

  • Upon the completion of the microarray scan, customers will receive raw scan files on DVDs. These files include: CEL file (cell intensity), .DAT file (raw image), and ARR file (sample information).

  • Affymetrix provides a free software- Transcriptome Analysis Console, which analyzes most gene expression and alternative splicing arrays. This software must be downloaded on a 64-bit PC.

  • Affymetrix Software User Manuals
Steps to analyze CEL files:
  1. Download the Affymetrix Expression Console Software (64-bit version), using this link, http://www.affymetrix.com/estore/browse/products.jsp?productId=131414#1_1
  2. Import the CEL files and run appropriate analysis, depending on your array type.
  3. From the Tools option in the upper menu, open Transcriptome Analysis Console
  4. Import the CHP files that were created via the initial analysis in Gene Expression Console.
  5. Run Gene Level, Exon Level, or Alternative Splicing analysis, depending on your  array type.

 

Other software options:
Partek (accessible through CMGM)
GeneSifter
GeneSpring (accessible through CMGM)


 

Bioanalyzer QC : (back to top)

** Below are examples of the most common errors found in Bioanalyzer traces. This guide will help you understand your data better. If you have any questions or concerns about your data, please contact us at panbioanalyzer@stanford.edu.

** For further analysis of your samples, you can use the Agilent 2100 Expert Software to open the raw (.xad) data file.

RNA Assays

** General

DNA Assays

** General
** NGS samples


 

RNA Assays - (back to top)

Total RNA Integrity –   Spikes – Microparticles and/or air bubbles in the system

 

 

 

 

Broad Peaks – Sample contaminated with genomic DNA   Missing Peaks – Sample salt concentration is too high

 

 

 

 

Missing RNA Fragments – Sample salt concentration is too high   Wavy Baseline – contamination with genomic DNA

 

 

 

 

DNA Assays - (back to top)

** General --

Spikes –mechanical spikes are resulted from microparticles or air bubbles present in the system.  These spikes, in general, do not affect the quality and/or quantity of the sample.   Missing Upper Marker – may be caused by high salt concentration, ethanol contaminants and/or restriction enzymes present in the sample.

 

 

 


Missing Peaks – Sample salt concentration is too high   Baseline Dips – Sample concentration is too high

 

 

 


Peak Tailing – Sample concentration is too high
   


 

 

 

 

 

** NGS Samples -- (back to top)

High Molecular Weight DNA Contamination   Split Peaks – sample concentration is too high


 

 



Excess Adapter - Inefficient ligation due to too much input DNA or the use of incorrect ligation temperature (ligation is performed at 20-25°C)
  Primer Dimers and PCR Artifacts -



 

 

Bead Carried Over –
  Excess primer -


 

 


 

 


** References and Helpful Links - the summary above is based on the following resources. You can check them out for more details.