We used high-throughput tag-encoded 454 GS-FLX amplicon sequencing                 to assess fungal diversity in the selected plant communities. Amplicon libraries                 were performed using a combination of tagged primers designed for the variable ITS                 region, as recommended for the tag-encoded pyrosequencing method (Acosta-Martinez et                 al., 2008). Genomic DNA samples were amplified using the fungal-specific primer pair                 ITS1F (5'-XCTTGGTCATTTAGAGGAAGTAA) and ITS4 (5'-YxxxxTCCTCCGCTTATTGATATGC), where X                 and Y represent the two pyrosequencing primers (CCTATCCCCTGTGTGCCTTGGCAGTCTCAGT and                 CCATCTCATCCCTGCGTGTCTCCGACTCAGA) and xxxx represents the barcodes designed for                 sample identification. Four PCR reactions were performed per sample using the                 following temperature program: 95°C/2 min, 25 cycles of 95°C/30 s, 54°C/1 min,                 72°C/2 min, and 72°C/10 min. PCR products were pooled for each sample, purified and                 sequencing-adapters/tags were integrated via MegaPrimers (PCR). The amplicon length                 and concentration were estimated, and an equimolar mix of all amplicon libraries was                 used for pyrosequencing (with the ITS4 primer). Tagged samples were pooled and                 sequenced using the Genome Sequencer GS FLX 454 System (454 Life Sciences⁄Roche                 Applied Biosystems, Nutley, NJ, USA) at GATC Biotech AG (Konstanz, Germany) and the                 Leiden Genome Technology Center (Leiden, the Netherlands).