Target labeling for Agilent Arrays is done via a Fluorescent linear RNA amplification procedure.
This kit makes cyanine 3- or 5-labeled cDNA from microarrays using as little as 50 ng total RNA. The procedure consists of converting mRNA primed with an oligo dT-T7 primer into dscDNA with MMLV-RT, then generating amplified cRNA using T7 RNA Polymerase. The cRNA is reverse transcribed into cDNA and the cyanine dyes are incorporated.
This form must be filled out completely before we can accept any samples. Once this form has been filled out, please coordinate with Mila Angert to drop off samples.
The most critical aspect of any microarray experiment is the quality of the RNA. We require that the RNA be prepared using the RNeasy kit from Qiagen. This ensures high quality RNA, free from contaminants that can interfere with enzymatic reactions.
In addition to the tubes of RNA supplied for labeling, please provide a separate aliquot for Agilent Bioanalyzer analysis. Label these tubes with an A (denoting they are for Agilent analysis).
RNA quality will be checked at the facility. If low quality is suspected or the RNA fails to meet our criteria, the experiment will be stopped. Please contact Mila Angert if you have any questions or concerns about the itegrity of your RNA samples.
BIOGEM (BioMedical Genomics Microarray Facility)
University of California San Diego
9500 Gilman Drive
Tel: (858) 822-4231 or (858) 822-4506
Fax: (858) 822-6430
Email Contact: James Sprague