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A mean transcript sequence identity of 97% was found between ovine and bovine orthologs, which probably highlights the utility of the microarray technique with ovine samples [ 25].
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The "effective length" of each transcript which is the mean number of positions along the transcript sequence that a read may start (Li and Dewey 2011) was also computed using RSEM.
The normalized-FPKM (expected number of fragments per kilobase of transcript sequence per million base pairs sequenced) was used to quantify the transcript level in reads.
As a result of the assessment with CVG, we have derived the most comprehensive transcript sequence set of the Madagascar ground gecko by means of assembling individual libraries followed by clustering the assembled sequences based on their overall similarities.
Sequences were assembled into contigs of up to 3 kb in size (hsp90 transcript, Contig 8575, See Additional file 4), but the mean contig length of 745 bp indicated that most of them cover only parts of the cognate transcript sequence.
These sequences were designed from exons 2 and 3 of the PTH transcript sequence deposited as NM_000315 in GenBank.
This means that the overlapping EST sequences were aligned and joined in an as complete as possible representation of the corresponding transcript sequence (which is here termed 'contig'contig
Primer sequences for the transcripts examined are found in Table 2. Specific transcript levels were determined through comparison to a standard curve generated from the PCR amplification of template dilutions, and normalized to 18S ribosomal RNA and presented as mean transcript levels expressed as %18S RNA.
To distinguish between valid transcript sequence and spuriously assembled sequence we compared the post-clustering set of contigs to multiple Dipteran insect transcriptomes, searched for open reading frames and compared translated protein sequences to functional protein domain databases as a means to identify contigs with bioinformatic associations with other species.
The mean transcript length is 564 nucleotides.
The transcript sequence database consisted of contigs.
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