Tri6 auto-regulates its own expression in nutrient-rich conditions.
A) Arrangement of the Tri6 gene and location of Tri6 primers used in the RT-qPCR analysis of wildtype (Wt) and transgenic strains. The solid vertical box indicates the location of Tri6-ORF primers in the coding region of Tri6 (Filled horizontal box) of the wildtype and the Tri6 over-expressing transgenic strains (tri6ΔTri6). The dotted vertical box indicates the location of Tri6 5′ UTR primers in the wildtype (Wt), the Tri6 mutant (tri6Δand the Tri6 over-expressing transgenic strains (tri6ΔTri6). The location of the Hygromycin gene within the Tri6 coding region of the tri6Δstrain (tri6Δis indicated by the striped horizontal box. Gpd indicates the promoter used to over-expressTri6 . The solid horizontal lines indicate 5′ and 3′ flanking regions of the Tri6 gene. B) The quantitative real-time PCR (RT-qPCR) analysis of Tri genes in wildtype, tri6Δand the tri6ΔTri6 strains grown in nutrient-rich conditions. RT-qPCR reactions were performed in triplicates using Applied Biosystems Power SYBR Green kit and the Applied Biosystems Step One Plus Real-Time PCR System. A list of qPCR primers for all the Tri genes is listed in the Table S2. The β-tubulin gene (FGSG_09530) was used as the internal control and the data was imported and Relative quantity (RQ) was derived by the Relative standard method included in the StepOne 2.1 software. C) Identical to B), except the internal control used was Gapdh (FGSG_06257). The figures are representative of two independent biological replicates.