MacVector 18.1 and the new InterProScan functional domain analysis tool

MacVector allows you to do functional domain analysis on your protein sequence using the InterProScan service. InterPro contains multiple databases of protein families, domains and motifs and InterProScan will submit a protein sequence to a search of these databases. It will also do extra analysis such as transmembrane region analysis using TMHMM and other tools.MacVector will submit your protein sequence to an InterProScan search and allows you to permanently annotate results directly back to your sequence.

However, the InterProScan service is undergoing changes which means that this tool now has limited functionality. You can still submit sequences for analysis, and you will be able to view the results. However, the graphical interface is now not functional and does not allow you to directly annotate the results back to your sequence.

There is a workaround as MacVector does allow you to annotate protein sequences with GFF files using IMPORT FEATURES. GFF, along with BED and GFT, is a standard format for storing protein/DNA annotation.

However, we have been hard at work at replacing MacVector’s InterProScan tool and are pleased to announce that the new tool is available in MacVector 18.1. MacVector 18.1 was released in February 2021 but up to now has not been available via the inline updater. MacVector 18.1.3 is now the current release and contains the all new InterProScan tool. MacVector 18.1.3 is now available for online updating within MacVector and you should be prompted to upgrade shortly.

We’ve not just adpated the tool for the backend changes but made it better. It’s now got a similar interface to MacVector’s Scan DNA For.. tool. Scan DNA For automatically displays restriction sites, missing common features, primer binding sites and putative open reading frames directly on your sequence and allows you to permanently annotate them.

With the new InterProScan tool you will submit your protein sequence in the same way to the InterProScan service using DATABASE | FUNCTIONAL DOMAIN ANALYSIS (InterPro). However, when the results come back they will be presented on your existing sequence’s Results Window.

InterProScanFor clarity this sequence, Sars-COV–2 Spike Protein, had all existing features removed.

If you hover your mouse cursor over each domain then you will see a detailed list of that domain’s database entry.

InterProScanHoverHover over a result to see the database entry

To permanently annotate a domain to your sequence, use the context menu by right clicking and choose CREATE DOMAIN FEATURE.

InterProScan ContextMenu

Do remember that many tools within MacVector use Context menus that are available with a “right click”.

If you have a maintenance contract that was active on 1st February, 2021, then you can install MacVector 18.1. You will be prompted to upgrade in due course. However, if you have turned off online updates, then you can go to MACVECTOR | CHECK FOR UPDATES.. to upgrade or download the full installer.

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MacVectorTip: Context-sensitive Menus in MacVector

Although Apple are well known (notorious?) for always providing mice with only a single obvious button, in reality the Mac interface from early versions of MacOS all the way to macOS Big Sur, plus many Mac apps, have always used right click menus (or more accurately “context sensitive menus”) to provide extra functionality.

MacVector is no exception and there are a lot of shortcuts and tools that can be accessed using context-sensitive menus. You can access these by either right-clicking in a window, or by [ctrl] clicking if you don’t have a right-clickable mouse/trackpad. For the most part, it doesn’t matter where in each window you right-click – the same menu will appear. However, the menus may be slightly different depending on items you have selected in the current tab.

Here’s an overview of many menus that you can see throughout the various sequence editors in MacVector. At the end you will find some very useful tools for Assembly Projects and Align to Reference. But let’s first look at the different tabs of the single sequence editor.

Editor tab

Screenshot 2021 04 28 at 16 09 52

The menu shown does vary depending on whether then topology is linear or circular (Set Origin), whether the origin is non zero (Reset Origin to 1) and whether you have some sequence selected (Add to Feature).

Create Feature – creates a feature from the current selection.

Add To Feature – lets you add the current selection as an additional segment to an existing feature.

Set Origin – for linear sequences, lets you change the numbering origin to a specific positive or negative value.

Reset Origin to 1 – for linear sequences, resets the numbering origin.

Map tab

Screenshot 2021 04 28 at 16 12 36

Again this context menu is variable. Its content will change based on the object(s) you currently have selected. In this case, it was a restriction enzyme site (Add To Gel).

Hide Symbol – hides the currently selected graphical object.

Add to Gel – for Restriction Enzymes, adds the current molecule cut with the enzyme to the currently open Agarose Gel window. Will create a new gel if necessary.

Create misc_feature Feature – create a new feature based on the selection. This will vary depending upon the selected graphic object. For an RE Site it’s a misc_feature, for an open reading frame, it will be a CDS.

Set Circular Origin – for circular molecules, sets the “12 o’clock” position to the selected restriction site.

Zoom to Sequence – “zooms” the display to the current selection.

Fit to Window – equivalent to double-clicking in the window to reset the scale to match the current window size.

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Edit – edits the selected feature.

Delete – deletes the selected feature.

Create – creates a new feature with the current selection in the Editor tab.

Join – if you have two or more features selected, this will join them into a single segmented feature.

Duplicate – duplicates the selected feature.

/protein_id= – this item varies depending on the feature selected. In this case it opens a browser and attempts to find the protein with that specific ID in the NCBI database.

Results window

Screenshot 2021 04 28 at 16 01 09

In Results windows where you can annotate the results back to the original sequence, then right clicking allows you to directly annotate that results. Such analysis tools include, InterProScan, Restriction Enzyme Analysis, ORF analysis, Primer design, and more.

Hide All Results – hides all of the result graphics.

Create XXX Feature – annotates that result to the orignal sequence. The feature created depends on the analysis that has been run.

Zoom to Sequence – “zooms” the display to the current selection.

Fit to Window – equivalent to double-clicking in the window to reset the scale to match the current window size.

The Align to Reference and Contig Editors

The Align to Reference and Contig Editors are very similar, except the Align to Reference Editor has both a reference sequence AND a consensus sequence whereas the Contig Editor just has a consensus sequence. Both have an extensive context sensitive menu. Lets look at the Contig Editor first – this is the Editor you use to view contigs generated by phrap, velvet or SPAdes in the Assembly Project window.

Screenshot 2021 04 28 at 16 15 02

Export Consensus with Gaps – saves the consensus as a MacVector .nucl sequence, including any gaps.

Export Consensus without Gaps – saves the consensus as a MacVector .nucl sequence, but removes the gaps. This is the more common function you would typically use for saving the consensus. This is a shortcut to the File | Export Consensus As… function.

Export Selected Reads as FASTA – saves the currently selected reads into a single fasta formatted file.

Export Selected Reads as FASTQ – saves the currently selected reads into a single fastq formatted file. This and the FASTA version are shortcuts to the File | Export Selected Reads As... menu item.

Select Matching Pairs – if you have assembled paired reads, and if the names of the reads follow one of the typical naming conventions, e.g. appending /1 and /2 to the read names, or having “1” or “2” in the description, this will select the appropriate matching pair.

Select Overlapping Reads Containing Selected Sequence – one of the most powerful tools in the Contig Editor. Suppose you have a SNP, or a variant base in a repeat sequence. Select the residue(s) and this command will select all of the other reads that have those residues at that position. In conjunction with Select Matching Pairs, this is a great way of finding, selecting, then exporting pairs of reads representing specific SNPs or repeats in an assembly.

Circularize Consensus – None of the assembler algorithms automatically detect circular sequences. But MacVector will automatically look at the ends of contigs and if there is a significant overlap, will offer to create a new circular sequence using this command.

Here’s the menu invoked from the Align to Reference Editor.

Screenshot 2021 04 28 at 16 19 30

Most of the menu items are the same as for the Contig Editor, but there are a few additions that are greyed out in the Contig Editor

Align Selected Reads – a shortcut to align just those reads you have selected

Delete Selected Reads – does what it says

Reset (un-align) Selected Reads – the only way of actually reverting a read to its unaligned state.

Extend Reference with Selected Read – if you have a linear reference and a read extends beyond the left or right end of the sequence, this lets you extend the reference with the contents of the read. This is a great tool for building up an extended reference to create e.g. a patch sequence to use in subsequent assemblies to join together two contigs using reads you have isolated using the other Align to Reference tools.

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Make the most of your Apple Silicon Mac with MacVector 18.1

We are very pleased to announce that MacVector 18.1 is now available to download. MacVector 18.1 is a Universal Binary application, which means it runs natively on both Apple Silicon M1 Macs and Intel Macs. MacVector 18.1 matches the “Big Sur” look and feel. …and for the first time in many, many years the MacVector icon has changed to match the square look of macOS Big Sur icons.

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MacVector 18.1 is supported on macOS Sierra to macOS Big Sur.

BigSurMV18 1

But it’s not just how it looks. We ran some benchmarks to see how much faster MacVector now runs on an Apple Silicon MacBook Pro. We compared this against MacVector 18.0, which runs using Rosetta2 emulation. In some cases you can see that the native Apple Silcon MacVector 18.1 runs 200% faster than the emulated MacVector 18.0. That’s an impressive speed increase!

M1 BenchmarksTable
MacVector 18.0 (X86_64) vs MacVector 18.1 (ARM64) (h:mm:ss).

Here are other new features in MacVector 18.1 and earlier releases.

The release of macOS Big Sur together with Apple Silicon Macs are a major Apple milestone and signify the end of OS X and the coming of macOS 11. At MacVector we are proud that MacVector 18.1 is fully compatible with macOS Big Sur and will run natively on Apple Silicon Macs!


How to upgrade to MacVector 18.1

If you have a maintenance contract that was active on 1st February, 2021, then you can install MacVector 18.1. However, because this release is so close after the release of MacVector 18.0 and the universal binary is a major change, then you must install it manually. MacVector 18.1 will not be available through the automatic in app updater for a few months. MacVector 18.1 is also no longer supported on OS X 10.10 El Capitan. You must be running macOS Sierra to macOS Big Sur.

If you have an older version of MacVector then download the trial and request an upgrade quote.

Even if you have downloaded the trial in the past then downloading a new trial will give you a fresh 21 days to evaluate MacVector.

When a trial license expires it becomes MacVector Free. So if you decide against upgrading then you can just delete the trial license and easily go back to your current version. It’s risk free as MacVector files are backwards compatible.

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MacVectorTip: Viewing genotype changes in Align to Reference assemblies

The latest releases of MacVector, MacVector 18.0.1 (Intel) and MacVector 18.1.1 (Intel and Apple Silicon) have some tweaks to the output of the SNPs tab in the Align to Reference assembly window. The genotypes of any SNP changes now follow a consistent standard, and short deletions are also reported. If the region containing the nucleotide change(s) is within an annotated CDS feature, then the genotype of the amino acid change is also reported. Here’s an example of some of the common SARS-CoV–2 variant “S” genes aligned against the reference Wuhan-HU–1 genome;

Align2Ref GenotypeChanges

Here you can clearly see the two characteristic deletions in the B.1.1.7 UK variant – H69_V70del and Y144del along with the two amino acid changes that are believed to be critical for the increased transmission rate – N501Y and D614G. The B.1.135 South Africa variant also contains the N501Y and D614G changes, but does not have the two deletions.

MacVector 18.1 was released at the end of February and is an Universal Binary which runs on both Intel and Apple Silicon Macs. Since it follows so closely after the release of MacVector 18.0 it is not yet available through the online updater. However, if you are running an M1 Mac then you can download the installer.

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MacVector on Apple Silicon: benchmarks

MacVector 18.1 has just been released. MacVector 18.1 is a Universal Binary application, which means it runs natively on both Apple Silicon M1 Macs and Intel Macs. MacVector 18.1 also has graphical changes for a more authentic Big Sur look and feel. …and for the first time in many, many years the MacVector icon has changed to match the square look of macOS Big Sur icons.

Download the installer.

BigSurMV18 1

But it’s not just how it looks. It’s how well it runs. So we ran some benchmarks on the M1 Mac. We used an Apple Silicon MacBook Pro with 16GB of RAM. We ran using MacVector 18.0, which runs using Rosetta2 emulation (X86_64) and using MacVector 18.1 which runs natively on Apple Silicon (also known as ARM64).

For the benchmarks we used the following jobs:

  • A dotplot of a pair of E.coli genomes
  • A multiple sequence alignment of 16 mitochondria genomes (“Sample Files/Mammalian mtDNA genomes – DNA.msan”) with Clustalw, T-Coffee and Muscle.
  • Using Align to Reference to map sequencing reads against a subset of the L.paracasei genome (sequence are in “Tutorial Files/Contig Assembly/NextGen”)
  • Align to Folder of a section of the Campylobacter jejuni genome against a pair of compressed FASTQ files containing 750,000 pairs of reads.
  • Here are the benchmarks (times are in h:mm:ss):

    M1 BenchmarksTable

    In some cases you can see that the native Apple Silcon MacVector 18.1 runs 200% faster than the emulated MacVector 18.0. That’s an impressive speed increase!

    We also compared a Intel iMac* with the M1 MacBook Pro. Here we used Align To Reference to map a dataset of paired reads in compressed FASTQ files to the full Campylobacter jejuni genome.

    The iMac could map reads at 19,500 reads per minute. But the M1 Macbook Pro rattled through those reads at 39,400 per minute. The small 13“ ”consumer level" laptop runs faster than a considerably more expensive Desktop Mac! Wow!

    MacVector 18.1 will be released shortly.

    *iMac (Retina 5K, 27-inch, Late 2014) with a 4 GHz Intel Core i7, 32 GB RAM and a Fusion drive 12 TB.

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    CRISPR tools in MacVector 18

    If you are using CRISPR editing techniques then there are two useful functions in MacVector for you:

    • MacVector will scan Nucleic Acid sequences for Protospacer Adjacent Motifs (PAM) associated with the CRISPR Cas9 and related enzymes cleavage and modification functions.

    • MacVector’s Analyze | Align To Reference… tool is ideal for screening reads for the short insertions, deletions or substitutions resulting from CRISPR experiments. See our blog for more information.

    Finding CRISPR Cas9 PAM sites.

    • Open your sequence.
    • Choose** Analyze | CRISPR PAM sites…**
    • click on the OK.

    You can also use Scan DNA FOR.. tool to automatically show sites on all sequences.

    CloningClipboard
    CloningClipboard
    • Choose MACVECTOR | PREFERENCES | SCAN DNA
    • Choose the CRISPR PAM tab
    • Toggle Show CRISPR PAM sites on

    Screening sequences for CRISPR introduced INDELS

    • Open your reference sequence.
    • Choose** Analyze | Align To Reference…**
    • click on the Add Seqs toolbar button to add reads from different clones/experiments.
    Scan DNA For..CRISPR sites
    Scan DNA For..CRISPR sites
    • click on Align
    • Choose CRISPR INDEL DETECTION
    • Click OK to align the reads against the reference.
    • When the job has finished click VIEW to see the results.
    Analyzing CRISPR INDELS
    Analyzing CRISPR INDELS
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    Using the RE Picker to display restriction enzyme sites in your insert that do not cut your vector.

    It’s generally useful with restriction enzyme based cloning to know what restriction enzymes will cut your vector and not cut your insert, or vice versa.

    There are a few ways to accomplish this with MacVector, but a useful way is to prepare a file of enzymes that do not cut your vector. This is especially useful if you want to repeat this for multiple inserts with the same vector.

    You can quickly do this with the RE Picker. The RE Picker shows an interactive list of restriction enzymes. Only those that are shown in the table and checked are displayed in the Map tab. However, you can change the criteria to only show enzymes that do NOT cut.

    • Open up your vector and switch to the MAP tab.
    • Click on the RE Picker button again to show the RE Picker window.
    • Click SELECT ALL to select all enzymes
    • Slide both the left and right sliders off the CUTS control all the way to the left.

    The Cuts label should now indicate “0”. The enzymes now visible in the RE Picker are all those in the default restriction enzyme file that do NOT cut the target molecule.

    • Click SAVE CURRENT SET OF ENZYMES and choose a relevant name e.g. [VectorName-NoCutters).renz.
    • Open up your insert sequence.
    • Click RE Picker
    • Click SET ENZYME FILE and choose your NonCutters file.
    • Make sure the CUTS control is set appropriately.
    • Click SELECT ALL to ensure all enzymes are selected and will be displayed.

    Only enzymes that cut your insert will now be displayed.

    REPicker NoCutters

    If you are designing a screen for clones after a ligation, then check out our Agarose Gel tool video

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    MacVector, Apple Silicon and macOS Big Sur compatibility

    UPDATE – March 8, 2021 – MacVector 18.1 is now released and can be downloaded.

    Over twenty years ago Apple called an end to the Classic Mac with the release of OS X, followed five years later by the transition from PowerPC to Intel. A major step forward in the Macintosh world.

    Earlier this year Apple announced the next stage in the Apple world, Apple Silicon Macs and macOS Big Sur. Signalling the end of OS X and heralding the start of macOS Big Sur.

    This week sees the Apple Silicon macs on sale and macOS Big Sur is being released today (Thursday 12th Nov 2020).

    We are proud to announce that our upcoming release MacVector 18 will run natively on both Intel and Apple Silicon Macs running macOS Big Sur.

    Here at MacVector we are proud that MacVector is a true Macintosh application. MacVector is a Mac native application and has always been designed for the Mac platform, rather than a cross platform application with the inevitable interface compromises. Right from the beginning MacVector was designed to take full advantage of the Mac’s easy to use interface to bring powerful sequence analysis tools right to your desktop. The MacVector development team always follow Apple’s Human Interface Guidelines so we know that anybody familiar with a Mac will quickly feel at home using MacVector. The Mac has always been easy to use, and any molecular biologist can start using MacVector and start designing primers, aligning sequences or making constructs straightaway.

    BigSur MacVector1 0 OS9 1990

    MacVector 18 on an Apple Silicon Mac running macOS Big Sur

    MacVector 1.0 running on a Classic Mac running Mac OS 9

    The Apple Silicon Macs will be more powerful but consume less power than the Intel Macs. Apple Silicon and macOS Big Sur are great steps forward for the Mac and we are pleased to be part of it.

    MacVector and macOS Big Sur

    As ever full information on the compatibility with macOS Big Sur will not be fully known until after its release on 12th November, 2020. However, our developers have been developing and testing on development releases of macOS Big Sur for many months now. Please check back to this post for current information.

    Current information is always available on the compatibility table.

    MacVector 17.5 (the current release): We will be waiting until after the release of macOS Big Sur later today (12th Nov 2020) before officially supporting macOS Big Sur with the release of MacVector 17.5.6 Tests show that MacVector 17.5.5 seems to run properly. However, from experience we invariably find some minor bugs after the official release. Expect MacVector 17.5.6 later next week.

    MacVector 18.0: our upcoming release will of course be fully supported on OS X El Capitan to macOS Big Sur. As well as macOS Big Sur support there’s a lot of new features to like in this release.

    MacVector 18.1: Our last release of 2020 will be a universal binary and will run natively on both Intel and Apple Silicon macs.

    Older versions:

    MacVector 17.0: initial testing indicate that MacVector 17.0 runs OK. However, it will not be officially supported.

    MacVector 16.0 and earlier versions: Our initial tests show that MacVector 16.0 and earlier will not run at all. These versions relied on an Apple library, that has been removed from macOS Big Sur. There is no possible workaround so if your license is only valid for MacVector 16 then either upgrade MacVector, or do not upgrade to macOS Big Sur.

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    Virtual Gene Cloning from NGS RNA-Seq Data

    The NCBI Sequence Read Archive (SRA) database is a huge resource of Next Generation Sequencing experimental data. Many groups and laboratories deposit data here that they have generated for their own specific projects that can be datamined for other unrelated projects with a minimum of effort.

    MacVector contains a number of powerful tools that can be used to extract and analyze specific sequences from large quantities of NGS data. We recently used these tools to clone the sequence of 19 distinct C2H2 Zinc Finger proteins from NGS RNA-Seq data prepared from root tissue of the Aloe vera plant.

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    The basic steps to do this were;

    • Use Align to Folder to find and extract all pairs of reads that could potentially encode the conserved QALGGH domain from C2H2 Zn Finger proteins
    • Assemble the reads using phrap, velvet and/or SPAdes to generate multiple contigs
    • Analyze contigs to identify and translate protein-coding ORFs
    • Extend contigs when required using additional rounds of Align to Folder, contig assembly and Align to Reference
    • Annotate proteins using the built-in InterProScan function
    • Align proteins using ClustalW and visualize the shared QALGGH domains

    The full tutorial is available as a PDF and the required data files are also available to download direct from the SRA.

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    Working From Home: An overview of primer design workflows in MacVector.

    Working from home?. We want to help familiarize you with the wide range of functionality in MacVector that you may never have used before. Here’s an overview of workflows for designing, testing, documenting and storing primers. You may not have a PCR machine on the kitchen table, but why not take the time to store all your lab’s regularly used primers in MacVector’s Primer Database.

    Amplifying a gene

    You can design a set of primers to amplify a gene in as little as three mouse clicks.

    1. Open your sequence.
    2. Open the MAP view, and click on a feature.
    3. Go to ANALYZE | PRIMER DESIGN/TEST(PAIRS).
    4. Click OK.

    Design a single targeted primer with a tail.

    QuickTest Primer tool gives great flexibility for designing primers with tails or mismatches.

    1. Select a 20 bp region around the location you want your primer to be.
    2. Click ANALYZE > QUICKTEST PRIMER.
    3. Slide the primer along your template until the oligo is optimal.
    4. Add a restriction site or mutation in the optional tail.
    5. Hover over a green (or red) putative restriction site to view the change to your sequence.
    6. Double click on that putative site to add to your primer. Watch how the coding is changed.
    7. Click Add to Database.. to save your primer.
    MacVector

    Test a pair of primers.

    You can map a pair of primers against your template to see how well they would amplify your target.

    1. Open your template sequence
    2. Go to ANALYZE > PRIMER DESIGN/TEST(PAIRS).
    3. For the left primer click USE THIS Primer
    4. Type or paste in your forward primer
    5. Repeat for the reverse primer
    6. Click TEST

    To save a primer from QT Primer to the Primer Database.

    1. Open ANALYZE > QUICKTEST PRIMER (INDIVIDUAL)
    2. Design your primer
    3. Click ADD TO…
    4. Give the primer a name and add a comment. Click OK

    To save a primer from Primer Design to the Primer Database.

    1. Open ANALYZE > PRIMER DESIGN/TEST (PAIRS)
    2. Design your primer pair
    3. In the spreadsheet right click on a primer
    4. Choose ADD PRIMER TO DATABASE
    5. Give the primer a name and add a comment. Click OK

    To use the Primer Database Search:

    1. Open your sequence
    2. Select ANALYZE > PRIMER DATABASE SEARCH
    3. Choose parameters and click OK

    Design a primer to match an existing primer from the primer database

    The Primer database allows you to store your own collection of primers. You can design new primers to match regularly used ones.

    1. Open your template sequence
    2. Switch to the Map tab and select the region you want to amplify.
    3. Go to ANALYZE | PRIMER DESIGN/TEST(PAIRS).
    4. For the left primer click USE THIS Primer
    5. Use the drop down menu to enter the existing primer from the primer database
    6. Click OK
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