title: LSU sequencing for aquatic fungal diversity
design: READS ARE MERGED. Primers LR22R-LR3 (Mueller et al., 2016, FEMS Microbiol Ecol). All libraries for each sequencing lane were demultiplexed using the Illumina bcl2fastq 1.8.4 software (folder 'RAW'). 1 or 2 mismatches or Ns were allowed in the barcode read when the barcode distances between all libraries on the lane allowed for it. Reads were sorted by amplicon inline barcodes: 1 mismatch was allowed per barcode; the barcode sequence was clipped from the sequence after sorting; reads with missing barcodes, one-sided barcodes or conflicting barcode pairs were discarded. Sequencing adapter remnants were clipped from all reads, Reads with final length less than 100 bases were discarded. Primer detection and clipping (folder 'PrimerClipped'): 3 mismatches were allowed per primer; pairs of primers (Fw-Rev or Rev-Fw) had to be present in the sequence fragments; if primer-dimers were detected, the outer primer copies were clipped from the sequence; the sequence fragments were turned into forward-reverse primer orientation after removing the primer sequences. Combination of forward and reverse reads was done using BBMerge 34.48.