FIG 5.

Lcc9 was downregulated in lcc9 silencing strains. (A, B) The laccase activity of four transformants of pYSK-lcc9 antisense 1 (A) and six transformants of pYSK-lcc9 antisense 2 (B) tested in separated coculture with Gongronella sp. w5 was found to be decreased, using ABTS as the substrate. R1-7, R1-15, R1-18, R2-6, R2-9, R2-11, R2-19, and R2-23 were positive lcc9 silencing strains represented by solid lines. pCcAde8 transformants R1-C and R2-C used as control strains are represented by dotted lines, as well as wild-type strains. (C) Native-PAGE of laccase activities of C. cinerea wild-type and of four C. cinerea lcc9 silencing strains from 48 h and 60 h separated coculture with Gongronella sp. w5 indicates downregulation in the lcc9 silencing strains. Equal amounts of culture supernatants (10 μl) were loaded onto the gels and activity-stained after gel-electrophoresis. (D) Quantification of lcc9 activity from the separated cocultures deduced from gels as shown in panel C. (E) lcc1, lcc5, and lcc9 laccase transcripts were shown by qRT-PCR analysis to be downregulated in the four lcc9 silencing strains. The lcc9 transcription level of wild-type strain at 36 h was set as the baseline. The data were analyzed using Student's t test (*, P < 0.05; **, P < 0.01; ***, P < 0.001). Data show mean ± SD, n = 3.