Parameters

Reference database (ref) mode

A database file of nucleotide sequences must be specified using the Reference Database (ref) option. The database may be in FASTA format. The reference database should include sequences that might appear as parents in the query set. These should be high-quality sequences that are believed to be free of chimeras. Errors in reference sequences will degrade detection accuracy and increase the number of false positives. Chimeras will not be detected if their parents (or sufficiently close relatives) are not present in the database.

The reference database should contain high-quality sequences that are believed to be chimera-free.

De novo mode

De novo chimera detection using the UCHIME algorithm. The input file must contain estimated amplicons with abundances specified by size annotations. In de novo mode, abundance skew is used to distinguish chimeras from parents. input should be estimated amplicon sequences with integer abundances specified using size annotations, e.g.:

>FQ23BBGZ5;size=23;

The minimum abundance skew is specified by the .abskew parameter, which defaults to 2.0 (because one round of PCR doubles the abundance). Abundance is a measure of how many amplicons with a given unique sequence were present in the sample after amplification by PCR. One way to estimate this is to sum the total number of reads in the cluster used to estimate the given amplicon sequence. UCHIME uses only ratios of abundances, so the absolute value does not matter. However, using the number of reads is a useful indicator.for example, a cluster containing one read is likely to be spurious. Amplicon sequences and abundances can be estimated using USEARCH, or by using another algorithm such as Chris Quince's PyroNoise or AmpliconNoise. When using de novo mode, sequences should be estimated amplicons from one sequencing run (strictly, one PCR amplification stage), otherwise abundances may not be directly comparable.

Inputs

Reference database mode

1. An input file containing the sequences in FASTA format.
2. A reference database file in FASTA format containing nucleotide sequences believed to be free of chimeras.

De novo mode

1. A FASTA file containing for each sequence estimated amplicons with abundances specified by size annotations, e.g. >FQ23BBGZ5;size=23; .

Output

This tool produced four output files two of which are hidden by default.

To view the hidden files: click on the cogwheel icon in the history panel and select 'Include Hidden Datasets'.

1. A FASTA file of predicted chimeras
2. A FASTA file of non-chimeras
3. (hidden) A human readable file of chimeric alignments
4. (hidden) A tab-separated file with the following 18 columns: