Frequently Asked Questions


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SeqMan NGen


» What file extensions are used for unassembled sequences? 
In SeqMan NGen 2.2 and earlier, unassembled sequences were saved in FASTA and QUAL formats (*.fas., *.qual). In SeqMan NGen 3.0 or later, these sequences are instead saved in FASTQ format (*.fastq).

Note that FASTQ files created with the SeqMan NGen wizard will have a *.fastq extension, while those created via a command line script will have a *.fas extension.
» Why can't I add a downloaded genome package to SeqMan NGen's "Add Template Files" dialog? 

Downloaded genome packages are saved on your computer as ZIP files, and must be extracted prior to use.  

To extract the genome package:

  • • On Macintosh:  Double-click on the ZIP file.  The files will be automatically extracted via the Archive Utility.
  • • On Windows Vista or Windows 7:  Double-click on the ZIP file.  In the ensuing Explorer window, click Extract all files from the top left.  Choose a location for the files and select Extract

To add the genome package to SeqMan NGen:

  1. 1. Open the SeqMan NGen wizard and follow the on-screen instructions.
  2. 2. In the "Add Template Files" dialog, select Add Genome Package.
  3. 3. Navigate to the extracted file, which has the extension *.genometemplate.
  4. 4. Click Open.
» I'm a Linux user with an Ubuntu operating system. Why doesn't SeqMan NGen appear on Ubuntu's "installed software" list? 

SeqMan NGen does not appear in the Ubuntu Software Center's Installed Software list.  This issue has been observed on Linux machines running Ubuntu 10.04, but may occur with other versions, as well.

The issue can be resolved using either of the methods below:

  • Perform at least one SeqMan NGen assembly prior to opening the Ubuntu Software Center.
  • Open the Synaptic Package Center by choosing SYSTEM > Administration > Synaptic Package Manager. Search for SeqMan NGen. After confirming the presence of SeqMan NGen, close the Package Manager and re-launch the Software Center.
  • After using either method, SeqMan NGen Linux should now be listed in the Software Center's Installed Software list.

    » In the Assembly Report, why is the “Export Aligned” value greater than the total number of sequences? 
    Since split reads are counted in each location where they align, it is possible for the value in the export aligned column (“Export_Aligned”) to exceed the total number of sequences (“NumSeqs”). The number of exported split reads can be viewed in the Assembly Report’s “Export_Split_Cnt” column.
    » My SeqMan NGen assembly used multiplexed data. Why is the “MID” column missing from the SeqMan Pro SNP Report? 
    In order for SeqMan Pro to display the MID column in the SNP Report, you must choose either the diploid bayesian or haploid bayesian method of SNP calculation prior to assembly in SeqMan NGen. This can be done from the following dialogs:
  • Assembly Options dialog > Advanced Assembly Options button > SNP tab > SNP calculation method drop-down menu = Diploid bayesian or Haploid bayesian.
  • Recalculate SNPs > SNP Options button > SNP calculation method drop-down menu = Diploid bayesian or Haploid bayesian.
  • If you instead choose the “simple percentage” method for SNP calculation from the drop-down menu, the MID column will be omitted from the SNP Report.
    » Why do assembly statistics vary from version 3.0 to 3.1? 

    When you perform the identical assembly in SeqMan NGen versions 3.0 and 3.1, some assembly statistics may appear “worse” in the newer version. This is actually due to an improvement in SeqMan NGen’s calculation algorithm. The criterion for calculating contig length is now more stringent and excludes positions that contain more than 50% gaps. In general, this causes the average contig length in version 3.1 to be shorter than in version 3.0. The statistics that may be affected by this improvement include:

  • Contigs to Reach Genome Length
  • Contig N50
  • Average Coverage
  • Average Lengths: Contigs
  • This change in the algorithm assures that you will get the most accurate assembly possible.

    » I'm a command-line user. When I type: sng [or smng, or xng] "path to script," why doesn't anything happen?  
    The application may not have been added to you environment variable PATH. After installing SeqMan NGen on a computer for the first time, command-line users may want to restart the computer before using the application. Restarting will add the application to your environment variable PATH.

    If you are unable to restart your computer, the following two options are available:
    • 1. Use the wizard to create and run the script.
    • 2. At the command-line, specify the path to the application followed by the path to the script.