Overview

The design of thousands of specific oligonucleotide probes or cDNA primer pairs is crucial to the success of any microarray project. The only choice available to the researcher thus far has been to launch a primer-design program designed to handle a single template in each run thousands of times, a tedious time-wasting task with limited assurance of the specificity of the resulting probes. Array Designer is the first and only software package that addresses this need by automatically selecting probes with optimal annealing characteristics and "BLASTing" them against any National Center for Biotechnology Information (NCBI) database to ensure specificity. Thus, thousands of specific probes can be designed in minutes.

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

Under an hour. After taking the 15-minute tour and exploring the menu items, I was able to start designing primers immediately.

Product Quality

Customizability

Not applicable.

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

Not applicable.

Ability to Import and Export in Different File Formats

Any sequence in GenBank, FASTA, or text format can be input. The program accepts multiple sequences in a single file in both FASTA and GenBank formats. The output is automatically loaded into a spreadsheet program.

Useful or Unusual Features

Great Time-Saver

Array Designer is the first and only program to design thousands of primers for making cDNA or oligonucleotide microarrays. Load all sequences of interest onto your local hard drive, select desired parameters, press the search button, and watch Array Designer rev up its search engine. Because Array Designer processes an average-length (around 2 kb) sequence in under a second, the entire primer-selection process for a microarray project can be reduced from weeks to days.

Searching for the Best

Although the substantial time savings is clearly the major advantage of the program, its power lies in the search algorithm at its heart. Array Designer looks at every position of the entire sequence, rates every possible oligo within the specified length range, and ranks each of them. Tm is calculated based on nearest-neighbor thermodynamic theory using SantaLucia values. Array Designer then evaluates secondary-structure stabilities of each oligonucleotide and outputs the best primers as close to the middle of the target range as possible using statistical optimization techniques.

cDNA Microarrays

Most labs prefer to use standard reaction conditions for all PCR analyses. Array Designer automatically selects primers for all the sequences with the same Tm or the target Ta (the annealing temperature).

To spot arrays with probes for all open reading frames (ORFs), you can use the ORF finder on the NCBI Web site and save the file containing the ORFs in FASTA format. To design primers to amplify almost the entire ORF, a special selection is provided to find the sense primers close to the 5' end of the template and the anti-sense primer near the 3' end of the template.

Probes for Oligonucleotide Microarrays

Synthesized oligonucleotide probes are used for detection of single-nucleotide polymorphisms (SNPs) as well as in gene-expression studies. Array Designer supports probe design for both needs.

For SNP detection, you can design probes that overlap the SNP sites by providing the sequence number of the SNP sites of interest in the input file. If you are using a primer-extension technique, probes can be designed with their 3' end one base upstream of the SNP site.

Usually, probes hybridizing near the 3' end of the gene are used for gene-expression studies. Probe length is around 50 to 70 nucleotides. Difficulty in designing probes guaranteed to bind uniquely to each gene of interest has led to the practice of using two to three probes for each gene to ensure the identity of each gene while sacrificing microarray real estate. Probes fulfilling these requirements can be designed in a single run by creating the appropriate input file.

BLAST

To assure the specificity of primers, each designed amplicon or probe can be submitted to the NCBI server for a BLAST search. Array Designer makes this tedious process convenient by batch processing a large number of searches. In addition, it provides an easy-to-use interface for submissions and for viewing results.

Versatile Output

After completing the search, Array Designer automatically launches the spreadsheet program, such as Excel or Lotus 123, available on your computer to show you the results. The output file is formatted using comma-delimited fields. Most synthesis companies will accept that format for a synthesis order. The data can be easily imported into internal databases such as Oracle or Access. The output file is also compatible with most spotting-robot software and can be used for downstream processing.

Limitations

The major limitation is that Array Designer requires the user to download the sequences of interest to the local hard drive instead of specifying the accession number. In addition, it does not accept the SNP site specification in dbSNP format. The vendor plans to overcome these limitations in the next version of the product.

Comparisons with Similar Software

Currently there are no other software packages for designing primers for microarrays.

Technical Support and Documentation

The ease of using Array Designer is due in part to the streaming multimedia tour available on the Web site. It provides a good introduction to the search concepts used by Array Designer and is an effective way to learn to use the program.

The online help and the manual are useful, covering every function in a concise manner. Technical support is freely available through email and by telephone. Support staff is helpful and knowledgeable.

Target Users

The program is useful to any microarray group designing custom microarrays, whether in academic institutions, industry, or core facilities.