Providing a stable methodological basis for comparing transcript abundance of developing embryos using microarrays.
High throughput methods deliver large amount of data serving to describe the physiological treatment that is being studied. In the case of microarrays, there would be a clear benefit to integrate the published data sets. However, the numerous methodological discrepancies between microarray platforms make this comparison impossible. This incompatibility is magnified when considering the peculiar context of transcript management in early embryogenesis. The total RNA content is known to profoundly fluctuate during development. In addition, the mRNA population is subjected to poly(A) tail shortening and elongating events, a characteristic of stored and recruited messengers. These intrinsic factors need to be considered when interpreting any transcript abundance profiles during early development. As a consequence, many methodological details affect microarray platform performances and prevent compatibility. In an effort to maximize our microarray platform performance, we determined the various sources of variation for every one of the main steps leading to the production of microarray data. The five main steps involved in sample preparation were evaluated, as well as conditions for post-hybridization validation by qRT-PCR. These determinations were essential for the implementation of standardized procedures for our Research Network but they can also provide insight into the compatibility issues that the microarray community is now facing.
Gilbert
Isabelle
I
Centre de recherche en biologie de la reproduction, Département des Sciences animales, Université Laval, Québec, Canada.
Scantland
Sara
S
Sylvestre
Eve-Lyne
EL
Dufort
Isabelle
I
Sirard
Marc-André
MA
Robert
Claude
C
D047108:Embryonic Development; D006801:Humans; D046228:Microarray Analysis / Q000379:methods*; D012313:RNA / Q000096:biosynthesis; D027724:Reproductive Techniques, Assisted; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
D012313:RNA