Effects of RNA degradation on gene expression analysis of human postmortem tissues.
The Affymetrix GeneChip platform was used to build a gene expression database of the normal human body. Postmortem human tissues represent a valuable source of biological materials for this type of study, but their use entails some delays before harvesting such tissues. We first evaluated the RNA quality obtained from tissues obtained 3-5 h postmortem and found variations that were both tissue and donor-dependent. RNAs extracted from brain regions were of higher quality than those obtained from the gut, while the cause of death was a significant factor in donor-dependent differences. To avoid these variables, we used rat duodenum to determine the effects of RNA degradation on the analysis of gene expression. Surprisingly, even samples exhibiting significant RNA degradation yielded robust gene expression results, comparable to those obtained using intact samples at a certain signal intensity cutoff. We extended these findings to our human expression database and obtained similar results, indicating that the Affymetrix platform, which is biased to the 3' end of transcripts for detection, can tolerate significant RNA degradation, while still yielding high quality expression data. Our resulting body index expression database is a valuable research tool. As examples of potential uses, we report novel expression sites for four potential therapeutic targets--CCL27, GPR22, GPR113 and GPR128--and as well as a set of thymus-specific genes, including three not previously associated with the thymus.
Lee
Jerry
J
Department of Molecular Medicine, Neurocrine Biosciences Inc., San Diego, California, USA.
Hever
Aniko
A
Willhite
Dorian
D
Zlotnik
Albert
A
Hevezi
Peter
P
D000818:Animals; D001921:Brain / Q000378:metabolism; D054425:Chemokine CCL27; D019742:Chemokines, CC / Q000235:genetics; D004386:Duodenum / Q000737:chemistry / Q000378:metabolism; D020869:Gene Expression Profiling; D006801:Humans; D008297:Male; D020411:Oligonucleotide Array Sequence Analysis; D009928:Organ Specificity; D012313:RNA; D020871:RNA Stability; D051381:Rats; D017207:Rats, Sprague-Dawley; D043562:Receptors, G-Protein-Coupled / Q000235:genetics; D013950:Thymus Gland / Q000378:metabolism
C492276:CCL27 protein, human; D054425:Chemokine CCL27; D019742:Chemokines, CC; D043562:Receptors, G-Protein-Coupled; D012313:RNA