To examine the consequences of this approach on the sequencing of highly expressed transcripts, the relative expression level of unigenes was estimated by calculating the transcript abundance expressed as reads per kilobase per million mapped reads (RPKM) [ 26].
Since the exon array generally contains 3 to 4 times as many probes per transcript as the U133 Plus 2 array and the probesets are distributed throughout the transcript cluster, the exon array may be more sensitive in detecting a subset of low expressing genes, at least in this data set.
The reverse primers were designed to target sites 150 nt downstream of the longest annotated transcript, as per the RefSeq annotation, in order to include downstream 3′end processing elements (Fig. 1).
A 1X solution of the double-stranded nuclease (DSN) enzyme was determined to be the appropriate concentration of enzyme to achieve a balance between under and over-digestion of transcripts as per the manufacturer's protocol recommendations.
TransDecoder was run using the TopHat/Cufflinks transcript assembly as per the instructions on the cited web page [ 56].
(See the Supplementary Material) RNA-Skim reports both Reads Per Kilobase per Million mapped reads (RPKM) and Transcripts Per Million as the relative abundance estimations of the transcripts, and both metrics are calculated by the way used in Sailfish (Patro et al., 2013).
Each transcript was annotated as per the best homologous protein and the corresponding annotation was assigned to it.
bHuman Genome Variation Society (HGVS) names are given for variants found with the coding region of a gene using the designation for transcript variant #1 as per the Ensembl/UCSC GRCh37/hg19 human genome assembly.
Temperature-dependent modulation of circadian phase has been related to changes in per transcript levels as well as PER and TIM protein levels [ 16, 21, 24, 26, 30- 34].
A significant correlation was found when considering nifH gene copy number per g of root (r = 0.60, P = 0.004, n = 18; Fig. 3E) but not per g of rhizosphere soil or per root system (Fig. 3C,G), as well as nifH transcript number per g of rhizosphere soil (r = 0.59, P = 0.004, n = 18; Fig. 3D) but not per g of root or per root system (Fig. 3F,H).
The revised text can be found in the second paragraph of the Results section and reads: "Next, we measured the abundance of these sequences as the number transcripts per million (TPM) within each library after the removal of rRNA reads.
