Novel subtractive transcription-based amplification of mRNA (STAR) method and its application in search of rare and differentially expressed genes in AD brains.
BACKGROUND
Alzheimer's disease (AD) is a complex disorder that involves multiple biological processes. Many genes implicated in these processes may be present in low abundance in the human brain. DNA microarray analysis identifies changed genes that are expressed at high or moderate levels. Complementary to this approach, we described here a novel technology designed specifically to isolate rare and novel genes previously undetectable by other methods. We have used this method to identify differentially expressed genes in brains affected by AD. Our method, termed Subtractive Transcription-based Amplification of mRNA (STAR), is a combination of subtractive RNA/DNA hybridization and RNA amplification, which allows the removal of non-differentially expressed transcripts and the linear amplification of the differentially expressed genes.


RESULTS
Using the STAR technology we have identified over 800 differentially expressed sequences in AD brains, both up- and down- regulated, compared to age-matched controls. Over 55% of the sequences represent genes of unknown function and roughly half of them were novel and rare discoveries in the human brain. The expression changes of nearly 80 unique genes were further confirmed by qRT-PCR and the association of additional genes with AD and/or neurodegeneration was established using an in-house literature mining tool (LitMiner).


CONCLUSIONS
The STAR process significantly amplifies unique and rare sequences relative to abundant housekeeping genes and, as a consequence, identifies genes not previously linked to AD. This method also offers new opportunities to study the subtle changes in gene expression that potentially contribute to the development and/or progression of AD.
Liu
Qing Yan
QY
Neurobiology Program, Institute for Biological Sciences, National Research Council of Canada, Ottawa, Ontario, K1A 0R6, Canada. qing_yan.liu@nrc.gc.ca
Sooknanan
Roy R
RR
Malek
Lawrence T
LT
Ribecco-Lutkiewicz
Maria
M
Lei
Joy X
JX
Shen
Hui
H
Lach
Boleslaw
B
Walker
P Roy
PR
Martin
Joel
J
Sikorska
Marianna
M
D020413:3' Untranslated Regions / Q000235:genetics; D000544:Alzheimer Disease / Q000235:genetics* / Q000378:metabolism; D001921:Brain / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation; D015723:Gene Library; D005796:Genes; D006801:Humans; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013382:Subtraction Technique; D014158:Transcription, Genetic
D020413:3' Untranslated Regions; D018076:DNA, Complementary; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger