The workflow sample, described below, takes FASTQ files with paired-end Illumina reads as input and process them as follows:

If you haven't used the workflow samples in UGENE before, look at the "How to Use Sample Workflows" section of the documentation.

Workflow Sample Location

The workflow sample "De novo Assemble Illumina PE Reads" can be found in the "NGS" section of the Workflow Designer samples.

Workflow Image

The opened workflow looks as follows:

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Workflow Wizard

The wizard has 4 pages.

  1. Input data: Illumina paired-end reads: On this page, files with Illumina paired-end reads must be set. 

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  2. Trimmomatic settings: The Trimmomatic parameters can be changed here.

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    To configure trimming steps use the following button:

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         <img src="/wiki/download/attachments/22062035/De novo Assemble Illumina PE Reads_3.jpg"/>
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    The following dialog will appear:

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         <img src="/wiki/download/attachments/22059547/Improve Reads with Trimmomatic Element.png"/>
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    Click the Add new step button and select a step. The following options are available:

    Each step has the own parameters:

    AVGQUAL

    This step drops a read if the average quality is below the specified level.

    Input the following values:

    CROP

    This step removes bases regardless of quality from the end of thread, so that the readhas maximally the specified length after this step has been performed. Steps performed after CROP might of course further shorten the read.

    Input the following values:

    HEADCROP

    This step removes the specified number of bases, regardless of quality, from the beginning of the read.

    Input the following values:

    ILLUMINACLIP

    This step is used to find and remove Illumina adapters.

    Trimmomatic first compares short sections of an adapter and a read. If they match enough, the entire alignment between the read and adapter is scored. For paired-end reads, the "palindrome" approach is also used to improve the result. See Trimmomatic manual for details.

    Input the following values:

    There are also two optional parameters for palindrome mode: Min adapter length and Keep both reads. Use the following dialog. To call the dialog press the Optional button.

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         <img src="/wiki/download/attachments/22059547/Improve Reads with Trimmomatic Element_1.jpg"/>
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    LEADING

    This step removes low-quality bases from the beginning. As long as a base has a value below this threshold the base is removed and the next base will be investigated.

    Input the following values:

    MAXINFO

    This step performs an adaptive quality trim, balancing the benefits of retaining longer reads against the costs of retaining bases with errors. See Trimmomatic manual for details.

    Input the following values:

    MINLEN

    This step removes reads that fall below the specified minimum length. If required, it should normally be after all other processing steps. Reads removed by this step will be counted and included in the "dropped reads" count.

    Input the following values:

    SLIDINGWINDOW

    This step performs a sliding window trimming, cutting once the average quality within the window falls below a threshold. By considering multiple bases, a single poor quality base will not cause the removal of high-quality data later in the read.

    Input the following values:

    TOPHRED33

    This step (re)encodes the quality part of the FASTQ file to base 33.

    TOPHRED64

    This step (re)encodes the quality part of the FASTQ file to base 64. 

    TRAILING

    This step removes low-quality bases from the end. As long as a base has a value below this threshold the base is removed and the next base (i.e. the preceding one) will be investigated. This approach can be used removing the special Illumina " low-quality segment" regions (which are marked with a quality score of 2), but SLIDINGWINDOW or MAXINFO are recommended instead.

    Input the following values:

    To remove a step use the Remove selected step button. The pink highlighting means the required parameter has not been set.

     

  3. SPAdes settings: Default SPAdes parameters can be changed here. 

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    The following parameters are available:

    Dataset type

    Select the input dataset type: standard isolate (the default value) or multiple displacement amplification (corresponds to --sc).

    Running mode

    By default, SPAdes performs both read error correction and assembly. You can select leave one of only (corresponds to --only-assembler, --only-error-correction).

    Error correction is performed using BayesHammer module in case of Illumina input reads andIonHammer in case of IonTorrent data. Note that you should not use error correction in case input reads do not have quality information(e.g. FASTA input files are provided).

    K-mers

    k-mer sizes (-k).

  4. Output Files Page: On this page, you can select an output directory:

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