Journal: BMC Genomics
Article Title: Gene discovery and transcript analyses in the corn smut pathogen Ustilago maydis: expressed sequence tag and genome sequence comparison
Figure Lengend Snippet: RT-PCR confirmation of antisense transcription. a) A graphic representation of um02794 transcription. The grey line represents the genomic sequence (middle), the blue arrow represents predicted gene structure, and the red (top) and green (bottom) arrows represent sense and anti-sense ESTs respectively. The range of the genome coordinates was included. b) Detecting antisense transcripts corresponding to um02794 via strand specific RT-PCR. In lanes 2 to 5 first strand synthesis was carried out on RNA of CM grown haploid cells. In lanes 6 to 9 first strand synthesis was carried out on RNA of MN grown haploid cells. First synthesis reactions of lanes 2 and 6 were prepared using sense strand specific primers; lanes 3 and 7 anti sense specific primers; lane 4 and 8 oligo dT and lanes 5 and 9 DEPC-treated water. Lane 10 used genomic DNA from U. maydis strain 521 and lane 11 used water a PCR template. Lane 1 and 12: Full Ranger DNA ladder.
Article Snippet: Reverse transcription was carried out on 5 μg of DNaseI treated RNA using Superscript III (Invitrogen) and oligo (dT)12–18 primers (Invitrogen).
Techniques: Reverse Transcription Polymerase Chain Reaction, Sequencing, Polymerase Chain Reaction