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Components of variance in transcriptomics based on electrophoretic separation of cDNA fragments (cDNA-AFLP)

dc.contributor.authorKarlovsky, Petr
dc.contributor.authorWeiberg, Arne
dc.date.accessioned2010-11-15T11:30:43Z
dc.date.accessioned2011-06-29T15:43:02Z
dc.date.available2010-11-15T11:30:43Z
dc.date.available2011-06-29T15:43:02Z
dc.date.issued2010
dc.identifier.urihttp://resolver.sub.uni-goettingen.de/purl?gs-1/5708
dc.description.abstract The sources of variance and errors in transcriptomics based on the electrophoretic separation of amplified cDNA fragments were investigated using cDNA-AFLP. Transcriptome profiles of the plant-pathogenic fungus Verticillium longisporum were generated by a standard cDNA-AFLP protocol followed by electrophoretic separation of amplified DNA fragments in flatbed polyacrylamide gels with fluorescence detection as well as by capillary electrophoresis (DNA sequencer). The total variance was partitioned into contributions of cDNA synthesis, adapter ligation, preamplification, amplification, and electrophoresis. Parameters of computer-aided peak recognition and matching were investigated and strategies improving matching success based on double passage with different signal intensity thresholds were developed. The overall quality of data was similar for cDNA-AFLP and microarray hybridization. Variance of cDNA-AFLP was independent of signal intensity while microarray data showed higher variance for low intensity signals. Capillary electrophoresis significantly reduced the number of wrongly matched and unmatched signals as compared to flatbed gels. These results are also likely to apply to related electrophoresis-based transcriptome analysis techniques such as mRNA differential display.
dc.format.mimetypeapplication/pdf
dc.language.isodeu
dc.publisherWiley-Blackwell
dc.rightsopenAccess
dc.rights.urihttp://goedoc.uni-goettingen.de/goescholar/License_Goedoc_2009_03.txt
dc.subject.ddc630
dc.titleComponents of variance in transcriptomics based on electrophoretic separation of cDNA fragments (cDNA-AFLP)
dc.typejournalArticle
dc.identifier.doi10.1002/elps.200800756
dc.type.versionsubmittedVersion
dc.bibliographicCitation.volume30
dc.bibliographicCitation.issue14
dc.bibliographicCitation.firstPage2549
dc.type.subtypejournalArticle
dc.identifier.pmid19588459
dc.description.statuspeerReviewed
dc.bibliographicCitation.journalElectrophoresis: Regular issue/8 May 09/In progress


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