Amplicon sequencing

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Amplicon sequencing and downstream metagenetic analysis

Often, Illumina amplicon sequencing is used to assess the diversity and composition of microbial populations. For many purposes, the entire downstream bioinformatic analysis may be carried out using only the Qiime pipeline. However, for unusual or very large data, some steps will not work to satisfaction. Hence, alternative bioinformatic programs may have to be found.

This wiki contains a quick overview over the typical steps included in the analysis of amplicon sequencing data. Also, some of the alternative programs are listed, together with reasons to use them, rather than Qiime.

If using Qiime, consult the documentation at . Qiime is installed on Abel; the default version is v1.5.0, but versions v1.8.0 and v1.9.1 are available as well. Load the default version of Qiime as

module load qiime

or the newest verion as

module load qiime/1.9.1

Quality control of fastq read files

Paired-end read merging

Removal of PCR primer sequences

Converting fastq files into fasta files

De-replication of reads

Clustering of reads

Removing singletons

Selecting representative sequences for clusters


Assigning taxonomical identifiers

Analysing OTU tables