To determine whether hph expression was responsible for cleistothecia production by UC1, cleistothecia production was tested using the strain UC26. UC26 is a derivative of UC1 in which the hph gene has been excised from the integrated T-DNA region by Cre-mediated recombination [21]. RNA levels of MAT1-1-1 and PPG1 were still increased in UC26 compared to G217B (Figure 3A, B). UC26 also still formed empty cleistothecia when paired with UH3 (Figure 1B), indicating Stem Cells inhibitor that hph expression is not necessary for cleistothecia production by UC1. Effects of T-DNA insertion on genes flanking site of this website integration Expression patterns of genes flanking the site of T-DNA integration

may have been altered in UC1 due to the insertion, and this might be responsible for the differences between UC1 and G217B. Effects of the site of T-DNA integration were analyzed in UC1 to investigate the cause of the differences between UC1 and G217B. It has previously been determined that the T-DNA selleck chemicals is integrated upstream of HCAG 08014 in the strain UC1 [21]. HCAG 08014 shares sequence similarity with the S. cerevisiae Bem1 protein, a scaffold protein involved in polarity and also in the pheromone response MAP kinase pathway. RNA levels of putative

Bem1 and of HCAG 08015, the two genes flanking the site of T-DNA integration, were analyzed. RNA levels of HCAG 08015 were undetectable in G217B and in UC1. RNA levels of BEM1 were increased in yeast phase UC1 and UC26 compared to G217B (Figure 3F). In mycelial phase organisms, when cleistothecia formation occurs, levels of BEM1 were increased in UC26 compared to G217B, but decreased in UC1 compared to G217B (Figure 3G). These results indicate that expression of the genes immediately flanking the T-DNA insertion site is not likely to be responsible for the ability of UC1 and UC26 to form empty cleistothecia. PRKACG Effects of T-DNA insertion site on cleistothecia formation To further explore the contribution of the site of T-DNA integration to the ability

of UC1 to form cleistothecia, additional strains were generated with the same T-DNA sequence integrated elsewhere in the genome. If the site of T-DNA integration plays a major role in UC1′s ability to form empty cleistothecia, then strains with the same T-DNA region integrated elsewhere in the genome would not be expected to form cleistothecia. If elements present within the T-DNA region are responsible for UC1′s ability to form empty cleistothecia, then strains with the same T-DNA region integrated elsewhere in the genome would still be able to form cleistothecia. To distinguish effects of the site of T-DNA integration on cleistothecia production from effects due to elements present within the T-DNA region itself, four additional strains were generated in the G217B background: ALT8, ALT13, ALT15, and ALT16.