Overview

Biotechnix 3d is an all-in-one Macintosh package for DNA- and protein-sequence analysis. This program allows the projection of 3-D structure projects using data from the Protein Data Bank (PDB). The complete package includes one year of free technical support, liquid crystal display (LCD) glasses, an infrared (IR) emitter, and connectors for 3-D visualization. Primary structure tools include DNA restriction analysis, PCR primer selection, base composition, DNA-to-protein translation, and open-reading-frame (ORF) identification and display. Secondary sequence analysis includes amino-acid composition, estimations of hydrophobicity, flexibility, local structure (e.g., helix, beta sheet), potential antigenicity, and estimates of accessible surface area. Biotechnix 3d is reputedly capable of active memory allocation, so there should be no need to quit the application to allocate more memory. It can also multitask, allowing slower analyses to run in the background while you continue using other aspects of the program.

How Long Did It Take to Learn to Use It Productively?

Installing the software was simple, but setting up the components required for 3-D viewing was less easy. One unexpected difficulty was that the overhang above the VGA port on the backs of the blue and white G3 and the G4 made it impossible to connect the VGA pass-through module without running to buy a $16 cable. That said, we were using prototype connectors, and what is being shipped with the product may be much easier to install.

One reviewer began to use this program within minutes of installing it. The other reviewer, who is more of a PC user, took a little longer, mostly because of lack of familiarity with the Macintosh interface (no left and right mouse buttons, selection of windows, etc.).

Product Quality

The general presentation of Biotechnix 3d is attractive and polished, as one might expect of Macintosh software. The pull-down menus are comprehensive and logical. A number of simple spelling errors occur that spoil presentation, e.g., "burried" for "buried" and "lenght" for "length." Biotechnix 3d crashed several times, although it was not very clear whether this was directly related to the program. Surprisingly, the CD version of the program sent to us by the developers proved to be significantly more unstable (crashed and required restarts every 10 to 15 minutes) than the demo version (which can be downloaded from the Web site). After reinstalling, the problem seemed to solve itself.

User Friendliness

The program is superficially user friendly; sequence-analysis protocols are stepwise after a fashion, and pull-down menus are, in general, easily utilized. However, the manual seems somewhat incomplete and "disconnected" from the program: it tells you what the program is supposed to be able to do without describing the precise steps required to achieve the desired task. Troubleshooting advice was sparse, and frustration set in at one stage when we attempted to follow stepwise instructions outlined in the manual that failed completely. On occasion, we found ourselves trying to follow impossible instructions using the online tutorials. The pull-down help menu for the 3-D viewer was surprisingly unhelpful. Many of the controls and viewing parameters are not readily obvious and require the user to refer to the manual. Once there, the explanations are incomplete, and not all of the items that are listed in the menu option are explained. There is no explanation of the color display options "temperature" or "structure." These presumably relate to the temperature factors and R values that must be included when the coordinates of the structures are deposited with the PDB (and, therefore, present in the downloaded file) and that are measures of the resolution for different regions of the structure.

Interface

Anyone used to the Macintosh will have no problem navigating through this program. Work is organized into individual projects, and the program generates a .PRJ file, which organizes Sequence, Alignment, and Trash folders for each project (you can directly access individual files through the Finder, so you don't have to go through the .PRJ file). A slight annoyance is that you must return to the .PRJ window before you can create a new sequence file. Organizationally, this makes sense, but when you just want to archive temporarily a small stretch of sequence or a DNA restriction fragment, it seems burdensome.

Some practical considerations when using the 3-D viewer might need to be worked out for individual users. We needed to work with a monitor in a dark room. The 3-D structures are displayed against a true black background similar to that obtained using graphics workstations. Thus, the glare created from ambient room lighting makes the structure projections difficult to see and can cause eyestrain. The developers suggest that higher monitor refresh rates may be easier on the eyes and allow you to work with ambient light. The refresh rate can be modified in the monitor control panel, where it is possible to select a resolution that gives you a higher refresh rate, for example, 85 Hz. By enlarging the window and zooming in on the molecule, you can also improve the 3-D quality as well as your visual comfort.

Intuitiveness of Design

The program is reasonably intuitive except for a few idiosyncrasies that take a bit of getting used to. For example, while the PCR primer-selection program finds plenty of primer pairs, it expresses the complementary (downstream) primer in the form 3'-5', which is contrary to standard notation, and not in a form that can be simply cut and pasted into another program or onto an oligo order form. We attempted to follow a recommendation to click/select the downstream primer, which would then be selected in the sequence in a 5'-3' format that we would be able to copy and paste onto an oligo order form, but we were unsuccessful. Clicking on the downstream primer highlights the equivalent sequence in the main sequence graphic, but when you copy/paste, you get the entire sequence as a screen snapshot, and when we tried the other option, which was to create a new sequence (necessitating, of course, a return to the .PRJ window), we could not paste anything into a newly created sequence file.

Customizability

None.

Ability to Program in Scripts, Add Extension Modules, etc.

None.

Ability to Import and Export in Different File Formats

One of the most useful and innovative elements of this program is displayed when trying to import files. While the program has little or no ability to import sequence files from other custom Macintosh sequence-analysis software, it has an exceedingly good interface with a host of sophisticated sequence databases and sequence-analysis sites accessible through the Internet. This keeps the program manageably small and ensures that the user doesn't have to keep updating software. We did a representative number of sequence analyses online (examples include PROSCAN [Prosite motif scan], SWISS-MODEL [sequence-based protein modeling], and HMMTOP [prediction of transmembrane regions and protein topology]). In some cases, going to the remote sites through Biotechnix allows you to browse back through your own hard drive to export the sequence - no confusion with formatting or cutting and pasting.

This feature is particularly valuable for people who regularly examine novel sequences that may not have been prepared in the FASTA format that is often required for online analysis. Importing sequence from GenBank was easy; we simply clicked and dragged the hyperlink from the GenBank page into the relevant project folder. The flat file display contains all of the associated GenBank annotations with the sequence. The same procedure worked equally well for Entrez sequence searches. This is some of the best point-and-click Internet interfacing that we have ever seen. However, the annotated information present in these files is not incorporated into the sequence as defined ORFs or other relevant sequence features. Biotechnix 3d is not the only program limited in this regard; it is a common frustration associated with sequence-analysis programs.

In contrast, exporting data in a useful format is extremely difficult if not impossible. This is the manual's sole comment on the subject: "Biotechnix 3d integrates seamlessly with your favorite word processor or presentation software. Results obtained from DNA and protein analysis can be simply pasted into a word processor or presentation document. These images can then be annotated and can form an integral part of your project report or presentation." The manual's assertion that you can simply cut and paste analyses into other programs is not exactly correct. We were unable to export any complete graphic format, e.g., hydrophobicity plots. They either came out as the plain sequence flat file, or the portion of the graphic that was on the screen at the time. Since many of these graphics are scrollable, simply selecting all and pasting into Word retrieved only the portion of the graphic that was visible in the window at that time. The exported graphic is effectively a snapshot of the screen, and would require further manipulation before using it for presentation. It is necessary to save the graphic as a Word picture before you can do any such manipulations (Word prompts you to do this when you attempt any alteration of the pasted image). This is also, as far as we can tell, the only way to save an analysis graphic. However, alignments of amino-acid sequences can be exported in a Clustal-style format that gives a rather unadorned and unformatted text file of the alignment. Unfortunately, such necessities as residue numbering are lost. Gentech developers are aware of these difficulties and hope to resolve them in future upgrades.

Useful or Unusual Features

Obviously, the most unusual feature of this program is the ability to project 3-D images of proteins, using the attached glasses and projector. We are fortunate to have a colleague working with us who uses 3-D modeling on a regular basis, and she was an enthusiastic participant in our trials. The program uses the file format with appropriate coordinates from the Protein Data Bank (via a direct link from within the program) and allows both stereoscopic and 3-D views to be displayed. The 3-D view can be rotated smoothly about all axes or set into a free-spinning mode using the mouse or on-screen directional controls. Our colleague was particularly struck by how smoothly the model could be manipulated and rotated without the "strobing effects" often seen when the computer is unsuccessfully trying to keep up with the mouse.

The location of specific amino-acid residues can be identified either while viewing the structure by clicking the mouse over the residue, or by going back to the sequence file and highlighting the residue in the original sequence file and then returning to the structural projection. Groups of adjacent residues can also be displayed (for instance, those forming a specific loop structure) by clicking the residues at either end of the protein section of interest or by highlighting the amino-acid segment of interest in the attached sequence file. In addition, distances between two residues within the structure can be measured, as can bond angles. This is very useful information if the user would like to determine whether the distance between two side chains is appropriate for H-bonding interactions, or to determine whether the conformation of a particular loop or beta strand is particularly strained.

Usefully, it is possible to show simultaneously via multiple windows a highlighted amino-acid sequence, the 3-D projection of the same region of the protein, a hydropathy plot, and associated analyses. This would be most convenient for researchers trying to design mutagenesis experiments or other manipulations.

We were somewhat surprised to find a sequence-analysis program that did nothing to disguise that it uses the old fashioned "2 + 4" rule of thumb when deriving possible primer pairs. Many primer-selection programs now offer a wide selection of primer-selection options incorporating all manner of energy and stability criteria, and indeed these factors are also incorporated into Biotechnix 3d when selecting primers longer than 20 mer. We subjected the primer selection component to a direct comparison test with a demo version of MacVector 7.0. The results were interesting, as the more stringent MacVector failed to recommend any pairs for a piece of test sequence, while Biotechnix 3d came up with several suggestions. Subsequent synthesis of the primer set and PCR was successful (leaving one reviewer positively disposed toward the program - his sequence and product). We shouldn't have been too surprised, it has been our experience that a PCR reaction can be much more forgiving than a selection formula.

Limitations

The 3-D display and analysis component is what differentiates this program from other personal computer sequence-analysis packages. However, it doesn't compare with the level of modeling that is currently achieved using state-of-the-art software such as Insight II running on an SGI or IBM RISC system/6000 workstation, and thus your ability to modify the model is quite restricted. We were not able to add significant data often not contained in a PDB submission, such as electron-density mapping, nor did we find a way to remove parts of the molecule to allow us to focus on a single side chain, or to examine parts of a model that might be of particular interest, such as the interface of two subunits or the chain segments that form the active site of an enzyme. This inability to focus on particular regions of the projection would certainly limit its usefulness for the analysis of tertiary structure homologies.

3-D viewing requires the use of a VGA monitor, so the latest Apple monitors (which use an Apple combined monitor/power/USB ADC connector) are not supported. Unfortunately, the 3-D component will not run on iMacs or PowerBooks. The IR emission/LCD glasses technology used for 3-D visualization is based on synchronization with the refresh rate of the screen. This technology simply does not work with LCD screens; the problem is not peculiar to Biotechnix 3d, but to any 3-D software using the same technology. PowerBooks will handle 3-D viewing if you connect them to an external Apple or other display. Unfortunately, this cannot be done with iMacs because they do not have any connector for an external display. This is disappointing because many Macintosh-oriented labs use iMacs as bench-top computers, but that is hardly the fault of the software developers.

One very tedious aspect of using the program was encountered when moving iteratively from the sequence file to the structural projection because the software "forgets" the orientation and display changes used to optimize viewing of the region of interest. A means of saving this information would certainly add to the ease with which this program could be used.

The restriction analysis section is rather clunky, although the graphics are colorful. We couldn't generate a simple "dense" double-strand sequence containing all the detected restriction sites. We were able to generate only a map consisting of a single line of sequence every one to two inches, so we could see only a few lines at a time. One reviewer was very disappointed with this aspect of the program. He regularly uses two Macintosh programs: GeneJockey, a small, comparatively ancient (1990) version of the sequence-analysis program, and DNA Strider (essentially homemade and really ancient, 1989) and finds them more agile and efficient for performing these kinds of analyses. The split-screen format of Biotechnix 3d with a box graphic indicating the relative position of restriction sites above, and straight numbered sequence below, looks attractive, but we found it more difficult to use than a simpler presentation with all the data clumped together.

On a practical level, the most serious weakness was the limited export capabilities and the inability to manipulate easily graphic data into a format that could be imported into programs often used for presentation. The manual and online tutorials still need more work to be completely comprehensive. This is particularly important for the 3-D viewing component because many users may have no previous experience with modeling packages.

Comparisons with Similar Software

One reviewer is most familiar with PC-based software such as Omiga and Sci-ed Central. In comparison with those programs, she would say that with the exception of Biotechnix 3d's Internet component, it seemed less flexible and lacked some useful modules, e.g., on-screen cloning, for the primary and secondary sequence analysis. David Carroll has previously used GeneJockey, DNA Strider, and various versions of MacVector (up to version 6.5). His view is that Biotechnix 3d falls somewhere between these programs. It is definitely a step up from the first two in terms of facilities and graphics, but it cannot compare with MacVector for easy customizability and the ability to export data. However, he found that Biotechnix 3d was more user-friendly than MacVector, and that the Internet interface was very impressive.

The 3-D component has great potential. The structure display, once we had figured out the hardware and furniture requirements, was impressively fluid and easy to visualize. Our modeling colleague spent a lot of time playing with the program, and although she had complaints, she was excited to be trying out a program that goes partway toward allowing her to do the kinds of analyses previously possible only with prohibitively expensive workstations and software.

Technical Support and Documentation

We were forced to use the online version of the manual, which can be a little uncomfortable to use, but a hardcopy version with quick-start guides will be shipped with the commercial product. We found the manual somewhat superficial, and it seems to have been constructed with a view to advertising the program rather than assisting in its use. A FAQ attempts to assist with troubleshooting. It is possible to email for technical support and assistance. The Biotechnix 3d Web site is currently under development, so potentially useful links from there may well become available. No phone-based help is available.

Target Users

Even without the 3-D facility, this is a reasonably useful general sequence-analysis program. The 3-D component is very promising and could well be useful to molecular biologists who are well versed with primary and secondary sequence analyses, PCR, etc., but who are inexperienced or who do not have access to the hardware and software for interactively viewing 3-D structures. The current limitations of Biotechnix 3d probably won't attract hard-core structural biologists who already have access to state-of-the-art modeling programs. Nevertheless, structural biologists who would like to use the implications of a structural analysis to study the underlying molecular biology of molecule would certainly benefit from a single program that allows them to move easily from DNA-coding regions, through the amino-acid sequence, to a 3-D model, and back. The ability to link structural information to sequence within a single file and software package will aid design of expression constructs or development of mutagenesis strategies. That it can run on a computer that costs less than $3,000 is a bonus.