Recombination hotspots are DNA sequences which enhance recombination around that region. HOT1 is one of the best-studied mitotic hotspots in yeast. HOT1 includes a RNA polymerase I (PolI) transcription promoter which is responsible for 35S ribosomal rRNA gene (rDNA) transcription. In a PolI defective mutant the HOT1 hotspot activity is abolished, therefore transcription of HOT1 is thought to be an important factor for the recombination stimulation. However, it is not clear whether the transcription itself or other pleiotropic phenotypes stimulates recombination. To investigate the role of transcription, we made a highly activated PolI transcription system in HOT1 by using a strain whose rDNA repeats are deleted (rdnDeltaDelta). In the rdnDeltaDelta strain, HOT1 transcription was increased about 14 times compared to wild-type. Recombination activity stimulated by HOT1 in this strain was also elevated, about 15 times, compared to wild-type. These results indicate that the level of PolI transcription in HOT1 determines efficiency of the recombination. Moreover, Fob1p, which is essential for both the recombination stimulation activity and transcription of HOT1, was dispensable in the rdnDeltaDelta strains. This suggests that Fob1p is functioning as a PolI transcriptional activator in the wild-type strain.