Linked Life Data

12074164

Source:http://linkedlifedata.com/resource/pubmed/id/12074164

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
6
pubmed:dateCreated
2002-6-20
pubmed:abstractText
Microarray expression analysis demands large amounts of RNA that are often not available. RNA amplification techniques have been developed to overcome this prcblem, but limited data are available regarding the reproducibility and maintenance of original transcript ratios. We optimized and validated two amplification techniques: a modified in vitro transcription for the linear amplification of 3 microg total RNA and a SMART PCR-based technique for the exponential amplification of 50 ng total RNA. To determine bias between transcript ratios, we compared the expression profiles in mouse testis versus spleen between the two amplification methods and a standard labeling protocol, using microarrays containing 4596 cDNAs spotted in duplicate. With each method, replicate hybridizations were highly reproducible. However, when comparing the amplification methods to standard labeling, correlation coefficients were lower. Twelve genes that exhibited inconsistent or contradictory expression ratios among the three methods were verified by quantitative RT-PCR. The amplification methods showed slightly more discrepancies in the expression ratios when compared to quantitative RT-PCR results but were more sensitive in terms of detecting expressed genes. In conclusion, although amplification methods introduce slight changes in the transcript ratios compared to standard labeling, they are highly reproducible. For small sample size, in vitro transcription is the preferred method, but one should never combine different labeling strategies within a single study.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
0736-6205
pubmed:author
pubmed:issnType
Print
pubmed:volume
32
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
1330-4, 1336, 1338, 1340
pubmed:dateRevised
2007-11-15
pubmed:meshHeading
pubmed:year
2002
pubmed:articleTitle
RNA amplification results in reproducible microarray data with slight ratio bias.
pubmed:affiliation
Hungarian Academy of Sciences, Szeged.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't