Five hundred ng of total RNA from each control and PPAR agonist-treated cell culture were separately reverse-transcribed into double-strand cDNA by the Moloney murine leukaemia virus reverse transcriptase and amplified for 2 h at 40°C using Quick Amplification Labeling Kit (Agilent).
In this study, total RNA of the different stages was separately reverse-transcribed and purified.
Two mRNA samples (experimental and reference) were separately reverse-transcribed, labeled, mixed, and hybridized in duplicate, with the dye assignments reversed in the second hybridization.
Discriminative motif discovery was performed by providing the regions around the methylated and unmethylated sites separately, and reversing the background and foreground sets to discover motifs enriched around both groups of sites.
Based on the feature of the stem-loop structure, a command line BLAST was performed to align each unique miRNA sequence with the sequences of the transcriptomic database, aligning forward and reverse strands separately by setting the parameter –S (1 for forward, 2 for reverse alignment).
These results are presented in figure 4. In each environment, we tracked the subpopulations of organisms with forward and reverse polymerases separately so that we could follow the population changes and polymerase rate evolution for each condition in the mixed case independently.
†Concentrations listed are for forward and reverse primers separately.
Parallel cell culture samples (biological replicates) were extracted and reverse transcribed separately.
RT-PCR was performed with RevertAid H Minus First Strand cDNA Synthesis Kit (Thermo Scientific TM, cat. No. K1631) using avidin and actin reverse primers, separately.
We calculated local dyad positioning scores in the forward and reverse strands, separately, and used the scores in the forward strand.
