We recently presented a webinar entitled “Best practices for sequence alignment in MegAlign Pro.”
You can view the recording of this webinar and others that we’ve presented on our Webinars page.
We asked for your questions before and during the webinar, and you delivered! We received dozens of questions from users around the world. We could only answer a handful of them during the live webinar, but we promised to email answers to each of you individually after the webinar. One attendee asked if we could post all Q&A online so everybody could see them. That sounded great to us, so here they are!
Topics Covered in this Article
Is MegAlign Pro free? Can I try it before buying?
MegAlign Pro is fully-supported commercial software. It is included as part of our Lasergene Molecular Biology package.
To buy the Molecular Biology package online or to compare package prices, visit our Pricing page. Discounted “Academic” prices pertain if you work or study at a university, government, or non-profit institution. To get a fully-functional 14-day free trial of Lasergene, click here.
Where can I find a User Guide? Is it in PDF format?
Our MegAlign Pro User Guide contains dozens of useful help topics, along with many tutorials on both pairwise and multiple alignment. Use the Download as PDF button under the table of contents to save it as a printable PDF.
MegAlign Pro and COVID-19 research
We received several questions regarding the use of MegAlign Pro in COVID-19 research. To see who is using Lasergene software for their COVID-19 research, check out our recent newsletters online (a recent example) or follow us on Twitter, Facebook or LinkedIn.
Support file formats and operating systems
- Can the program be used for amino acid sequences in addition to nucleic acid sequences?
Yes. MegAlign Pro supports DNA, RNA and protein sequences.
- What file formats are supported?
MegAlign Pro supports a wide variety of sequence and alignment file formats You can see a list of supported formats on our File Formats page.
- Can MegAlign Pro open files that were created in MegAlign?
Yes, MegAlign Pro can open any file made in classic MegAlign. In some cases, you may need to use the File > Import Alignment command to do this.
- Is MegAlign Pro 64-bit, and will it work with the latest release of Macintosh?
Yes, MegAlign Pro has always been 64-bit and works on all modern versions of Mac. Note that ‘classic’ MegAlign was 32-bit and would therefore not work on the Mac Catalina operating system. Since it had already been replaced by MegAlign Pro, MegAlign was therefore retired in 2019.
Layout / interface-related questions
- By default, MegAlign shows two views. Can I change the default layout to one panel showing sequences only?
Yes, learn how in the last table row on this page.
- I find scrolling back and forth along the alignment window to be slow and tedious. Are there tricks to help with this?
We agree! We recommend using our Viewport (see image) to navigate to the area of interest. Another thing that might help is splitting the Sequences view vertically so you can see two areas of interest at the same time.
- Classic’ MegAlign let me click on a residue or nucleotide to see its position in the sequence. How do I see the position in MegAlign Pro?
Click on the nucleotide or residue and view its information in the Details panel on the right. If the panel is not visible, use View > Details.
- How can I customize coloring of aligned residues?
This is done in the Sequence section of the Style panel.
- How do you edit information in the Details panel?
This information is not designed to be editable.
- Where are the alignment statistics?
Basic statistics can be found in the Details panel (View > Details) after performing an alignment. Pairwise distance statistics can be found in the Distance view.
Exporting data and images
- Can I export alignments to Microsoft Office applications like Word and PowerPoint?
You can export to PowerPoint (editable) or import as a .jpg and insert that into a Word document (non-editable). Information on both of these can be found here.
- Can I print alignment reports without exporting them first?
You can print the Sequence view directly using the File > Print command.
- How do I adjust the font size of MegAlign Pro reports?
In the Sequence and Overview sections of the Style panel (see image).
- How can I export an alignment into Excel to display Identical Residues and Similar Residues in two different background colors?
MegAlign Pro does not support exporting to Excel. However, you can export an image as follows: First, set up the Sequence view to show the color schemes. This is controlled using the Sequence and Multiple Alignment sections of the Style panel. Then use File > Export Image > Sequences.
- When doing protein alignments, what is the best way to export an image? / How do we export these alignment results for the purpose of presentation or for publication, especially 300 dpi tiff?
To learn how to export an image, see this topic. MegAlign Pro supports several high-resolution formats, as well as PowerPoint and PDF, but does not currently support .tiff.
Miscellaneous MegAlign Pro questions
- Can you align thousands of amino acid (aa) polypeptide sequences with a reference and know the most common amino acid at each position?
Yes. The Sequence Logo along the bottom of the Sequence view in MegAlign Pro can be used to assess the proportions of the three most abundant amino acids at a given position. (See it in the image near the top of this post). We plan for a more quantitative solution to be added to a future release.
- How do I make a consensus sequence using MegAlign Pro?
Simply perform a multiple alignment by clicking a button. The consensus sequence appears automatically at the top of the Sequences view. It can be exported using the commands File > Export Data > Aligned Consensus or File > Export Data > Aligned Consensus without Gaps.
- Is it possible to align a subregion?
Yes. After performing an alignment, select the subrange you would like to align and choose Align > Realign Subsequences. See this topic for detailed information.
- Can MegAlign pro group and quantify exact matches in an alignment of thousands of reads, as for amplicon sequencing? (So, % of Amplicon A, % of Amplicon B, C etc.)
Lasergene does not currently have a specific workflow for this, but the following steps might provide what you need. Perform a multiple alignment in MegAlign Pro; this will group identical sequences in the phylogenetic analysis. Then use the command Tree > Order Sequences Like Tree to help group identical and near-identical sequences in the other views.
- How can I align whole genomes for SARS-COV2 with other genomes and see the variation in just one window or figure?
We recommend performing a MAUVE alignment in MegAlign Pro for whole genome comparisons. If you are starting with sequencing data, first assemble your genome in SeqMan NGen, then export the consensus sequence for downstream analysis.
- How do I enter a large number of sequences in batches? Let’s say I am starting from a list of sequences in an Excel file.
A large list of sequences can be loaded into Lasergene software using standard methods. The tricky part of your question is getting the sequences in Excel into a compatible format. The data in the Excel file can look like anything so a conversion process would be different for each person or group. I find .fasta format very simple and accepted by most molecular biology applications. I would try to mimic .fasta, which has the following format, >Taxa Name on first line and sequence data on second line:
You can modify almost any Excel sequence data file into .fasta format by using a combination of the Concatenate function in Excel to add/remove characters (like adding the “>”), and Find/Replace function in a text editor using Extended Search Mode (“\r” can be used to insert a line break between Name and Sequence). Save as a text file and you are good to go. Once you know how to do this the process takes a few minutes.
- What is best algorithm to use in building a tree for Avian Influenza virus Strain H9N2? How do I select sequences to develop the phylogenetic tree?
A good resource for obtaining sequence data is NCBI. The best sequences to select would depend on your research question. We believe you are asking what the best phylogenetic method is for generating an Avian Influenza phylogeny. Phylogeneticists typically report more than one method. The two most commonly used and accepted are Maximum Likelihood and Bayesian Inference. We are currently working to include Maximum Likelihood In our MegAlign Pro application and will work on Bayesian Inference next.
When will version 17.1 be released? Will it include ____?
Many of you asked about the upcoming release and asked if specific new features would be included. We anticipate releasing Lasergene version 17.1 in June or July of 2020. This release of Lasergene is expected to include a number of new features in MegAlign Pro, including:
- The ability to reverse complement sequences
- Percent similarity (percent identity) reporting for pairwise alignments
- The ability to trim sequences and to trim and edit alignments
- Tree rooting and visual editing of trees
Questions pertaining to other DNASTAR applications and workflows
- How do I do transcriptomics in DNASTAR?
See our Transcriptomics Workflows for detailed information about de novo transcriptome assembly, RNA-Seq analysis, and ChIP-Seq analysis. These workflows require access to Lasergene Genomics.
- How can I find unique genes/regions in bacterial genome / COVID-19 genome assembly?
Lasergene Genomics supports genome assembly with or without a reference genome. If you are using a reference genome, you can do advanced variant analysis to find genes and SNPs of interest. See our Genomics workflows for more information.
- What application is used to do protein modelling and protein binding predictions?
Protein analysis can be performed with Lasergene Protein, which includes access Protean 3D and Nova applications for protein modeling. See our Protein Analysis workflows for more details.
- How to do cloning using DNASTAR software?
Lasergene supports a wide variety of cloning methods using SeqBuilder Pro. See our Automated virtual cloning page.
- How can I design a sequence and edit it?
See our Sequence editing and annotation page.
- How can I read the sequence? This function was there in EditSeq.
If you mean the ability to read the sequence as synthesized speech, this capability was only found in EditSeq. EditSeq was a 32-bit application that was retired in 2018. Its replacement, SeqBuilder Pro, does not support synthesized speech.
- I want to align ~5 sequences to check for a mutation from a site directed mutagenesis experiment. In the past, I used SeqBuilder to trim them, then aligned in MegAlign. What workflow should I follow now to easily see how my .abi files align with my template .sbd file?
Since you are using a reference sequence, this would be considered an assembly rather than an alignment. We therefore recommend starting in SeqMan Ultra. Click New Assembly on the left and then Sanger/ABI reference guided on the right to launch the SeqMan NGen assembly wizard. Follow the wizard steps to upload your .sbd and .abi files. Along the way, check Quality end trim to automatically trim poor-quality ends from the .abi sequences prior to assembly. Once assembly is complete, results will automatically open in SeqMan Ultra. In the Alignment view, bases that do not match the consensus are highlighted in yellow so you can easily check for the mutation.
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