We present evidence of a potentially serious source of error intrinsic
to all spotted cDNA microarrays that use I.M.A.GE. clones of expressed
sequence tags (ESTs). We found that a high proportion of these EST
sequences contain 5' end poly(dT) sequences that are remnants from the
oligo (dT)-primed reverse transcription of polyadenylated mRNA
templates used to generate EST cDNA for sequence clone
libraries. Analysis of expression data from two single-dye cDNA
microarray experiments showed that ESTs whose sequences contain
repeats of consecutive 5'-end dT residues appeared to be strongly
co-expressed, while expression data of all other sequences exhibited
no such pattern. Our anlaysis suggests that expression data from
sequences containing 5' poly(dT) tracts is more likely to be due to
systematic cross-hybridization of these poly(dT) tracts than true mRNA
co-expression. This indicates that existing data generated by cDNA
microarrays containing I.M.A.G.E. clone ESTs should be filtered to
remove expression data containing significant 5' poly(dT) tracts.
Keywords: cDNA Microarray, Expressed-Sequence Tag, EST,
Cross-hybridization, I.M.A.G.E. clone, systematic source of variation