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Microarray validation did prove the usefulness of the tool for further transcript quantification in Atlantic halibut.
About twenty ESTs spanning different functional groups were used for further transcript profiling analysis in our system during the later stages of mycorrhiza development.
Then, processed FLcDNAs/mRNAs with lengths of >29 bp after removal of the repeated regions were used for further transcript mapping.
We then removed the piRNAs that map to both gene transcripts and TEs, and used the remaining set of piRNAs for further transcript analysis.
We therefore elected to use the Trinity assembly to retain maximum information for further transcript analysis and the Trinity assembly is also available in Additional file 5.> To validate and annotate the assembled unigenes, the 16,798 unigenes generated by Trinity were subjected to BLASTX searches (E-value ≤ 1e-5) against public protein databases.
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A similar expression profile occurs for further transcripts of ATP synthase, arginine kinase and fumerase (found in Cluster B, Figure 3).
Ten transcripts lacked matching genomic sequences, for further ten transcripts the first exon cannot be found in the genomic database and for eight transcripts the second exon cannot be found.
Further, transcripts for almost all genes that were rhythmic at baseline failed to oscillate at the antiphase stage of forced desynchrony.
After signal processing, data normalization, and filtering, 8533 (83%) probes were kept for further analysis of transcript expression during early embryonic development and 8447 (82%) for analysis of differentially expressed transcripts between high and low quality embryos at 8CS.
For further analysis, the transcript levels were also examined by RTQ PCR, which revealed FOXO1 mRNA level was 15-fold highly expressed in KFr13Tx cells compared to KF28 cells.
TopHat-aligned read files were then entered into Cufflinks (version 1.2.1) software for further analyses, including transcript assembly, abundance estimation, and differential expression and regulation testing in RNA-Seq samples [ 42].
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