Five micrograms of total RNA were reverse-transcripted using a reverse transcription kit (Invitrogen Life Technologies, Carlsbad, CA, USA).
Ten of altogether twelve generally weakly expressed genes for isoprenoid biosynthesis, including three for the cytosolic MVA pathway, were amplified using a reverse transcription PCR approach with individually designed degenerate primers.
We report the first isolation of TPS gene from A. malaccensis using a reverse transcription approach.
Full-length chitosanase cDNA was obtained from total RNA by RT-PCR using a reverse transcription kit (TaKaRa).
cDNA was prepared from the bacterial RNA using a reverse transcription system (Fermentas).
RNA extraction was performed using Tri-reagent (Sigma) according to the manufacturer's protocol, and cDNA synthesis was carried out using a reverse transcription kit (Qiagen).
cDNA was synthesized by using a reverse transcription kit SuperScript III First-Strand Synthesis System for RT-PCR (Invitrogen) according to the manufacturer's recommendations.
RNA (2 µg) was treated with RNase-free DNase (Fermentas, USA) and then reverse transcribed using a reverse transcription system (Promega, USA).
In brief, 2 µg of total RNA from each sample was reverse transcribed for cDNA synthesis using a reverse transcription kit according to the manufacturer's protocol (Promega, Madison, WI).
Quality-tested total RNA was amplified using the RiboAmp RNA Amplification Kit (Arcturus, Moutainview, CA) and processed using a reverse transcription based method of label incorporation to yield labeled cDNA as previously described [8].
Reverse transcription (RT) was performed on 1 µg RNA at 60°C for 35 min using a Reverse Transcription kit (A3500, Promage) containing 0.5 µg random primers, 15 U AMV, 0.5 U RNasin RNase inhibitor.
