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Beacon Designerâ„¢ automates the design of real time primers and probes
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Beacon Designer™

Features

SYBR® Green Primer Design

  • Designs primers optimized for SYBR® Green PCR assays.

  • Beacon Designer™ designs highly specific SYBR® Green primers by BLAST searching sequences, automatically interpreting the results and then designing SYBR® Green primers by avoiding regions of significant homology.

  • The SYBR® Green primers designed can be BLAST searched against genomic databases available at NCBI. This helps verify the SYBR® Green primer design and visualize primer and amplicon specificity.

  • Beacon Designer™ identifies template structures and then automatically avoids them while designing SYBR® Green primers.

  • Displays the regions of homology and secondary structures for the sequence graphically in the Sequence View pane. This functionality helps visualize where the designed oligos are located with respect to both the regions.

  • Evaluates pre-designed primers and designs a compatible primer a given sense or anti-sense primer. It then proceeds to design optimal dual labeled probes for use with well proven SYBR® Green primers.

  • For SNP genotyping studies, designs SYBR® Green primers such that the SNP site at or towards t he 3' end position.

  • Calculates SYBR® Green primer Tm using nearest neighbor thermodynamic algorithm.
    Presents the design results in a ranked order.

  • Screens SYBR® Green primers for thermodynamic properties and secondary structures.

  • Avoids homologies with all other primers to reduce primer dimers in a multiplex reaction.

TaqMan® Probe Design

  • Designs optimal TaqMan® probes free of dimers, repeats and runs to ensure signal fidelity.

  • Designs wild and mutant TaqMan® probes for SNP identification for SNP genotyping assays
    Pre-designed primer sets can be evaluated and a compatible TaqMan® probe be designed.

  • Similarly, a pre-designed TaqMan® probe can be evaluated and compatible primers be designed. You can even import the primers designed in the SYBR® Green primer design mode and design a compatible probe.

  • Displays a graphical view of TaqMan® probe secondary structures.

  • Beacon Designer™ can design LNA™ probes. LNA™ substituted TaqMan® probes are shorter probes and have LNA™ bases that impart higher stability. You can specify the number of LNA™ bases that you wish to have in the probe. You can also specify their placement and the LNA free region on each end of the probe to design LNA™ spiked TaqMan® probes.

Read about how TaqMan® probes work!

Multiplex Real Time PCR Assay Design

  • Beacon Designer™ uses innovative proprietary algorithms to design optimal primer-probe sets for single tube multiplex assays.

  • Multiplex assays are supported in the TaqMan® design mode. You can multiplex up to five sequences.

  • Reference gene or housekeeping genes supported for normalization.

HRMA Primer Design

  • Designs mutation flanking HRMA primers such that they generate the shortest possible amplicon, for mutation detection.

  • The program identifies template structures and then automatically avoids them while designing HRMA primers, improving the PCR efficiency.

  • Beacon Designer™ checks the Tm, GC % and other parameters of the primers designed. Only the primers satisfying all the parameters of High Resolution Melting Analysis are reported.

  • Beacon Designer™ displays the sequence properties of both the wild and mutant amplicons along with their melting temperatures.

  • Facilitates verifying the specificity of the primers designed by primer BLAST.

  • Evaluates pre-designed proven primers and designs a compatible primer for a given sense or anti-sense HRMA primer.

MethyLight TaqMan® Design

  • Beacon Designer™ predicts CpG islands in accordance with Gardiner-Garden and Frommer guidelines. You can also specify the CpG regions manually in a sequence. The CpG islands are displayed in green in the Sequence view tab and the regions that are rich in GC are displayed in red.

  • Beacon Designer™ designs TaqMan® probes and methyl sensitive PCR primers for the specified CpG islands. You can also design suitable control primers and probes for MethyLight assays by designing them for unmethylated and untreated DNA sequences.

  • Primers and TaqMan® probes designed by Beacon Designer™ are highly specific as they are designed by avoiding regions of significant cross homology identified by automatically interpreting BLAST search results. The homologous regions to be avoided can be filtered on the basis of either the identity or the e value or both, the defaults for e value being 0 and that of identity being 100%.

  • The MethyLight assay involves treatment of genomic DNA with sodium bisulphite followed by an alkaline treatment. Beacon Designer™ displays the corresponding bisulphite treated strand when the genomic sequence is provided.

Read more about DNA Methylation.

Molecular Beacon Design

  • Automatically selects a stem of appropriate length for optimal beacon Tm.

  • Designs beacons free of dimers, repeats, and runs for increased signal strength.

  • Checks molecular beacons for cross homologies with all primers preventing competition in multiplex reactions.

  • Designs molecular beacons for detection of both wild and mutant alleles.

  • Pre-designed primer sets can be evaluated or primers for a pre-designed molecular probe can be constructed. Similarly, a pre-designed beacon can be evaluated or a beacon can be designed for a pre-designed primer set. This facilitates using SYBR® Green primers for beacon assays.

  • Displays designed beacon graphically along with its properties.

Read about molecular beacon technology.

NASBA® Assay Design

  • Beacon Designer™ can design molecular beacons for NASBA® assays for single template, multiplex or SNP genotyping assays.

  • Beacon Designer™ evaluates pre-designed primers and probes for NASBA® assays. While evaluating primers, Beacon Designer™ automatically adds a purine tag to P1 primer sequence when necessary.

FRET Probe Design

  • Optimal FRET Probes: Designs optimal FRET probes free of dimers, repeats and runs to ensure signal fidelity.

  • Allele Identification: Designs FRET probes for detection of both wild and mutant alleles.

  • Evaluate Pre-designed FRET assays: Pre-designed primer sets can be evaluated or primers for a pre-designed probe can be constructed. You can even import the primers designed in the SYBR® Green primer design mode.

  • Graphical View: Displays designed FRET probe graphically along with its properties.

Read about FRET probe technology.

Scorpions® Design

  • Beacon Designer™ supports Scorpions® chemistry for real time detection of a target sequence.

  • Beacon Designer™ designs Scorpions® primers that are free from dimers, runs, repeats and have user specified melting temperature ranges.

  • Beacon Designer™ automatically selects a stem of appropriate length for optimal Scorpions® Tm.

  • You can design specific Scorpions® primers by avoiding significant cross homologies that the target sequence has with the genome and by avoiding regions on the template that are susceptible to folding.

  • Beacon Designer™ displays the alternate structures of the Scorpions® designed graphically along with its properties.

  • Using Beacon Designer™, you can also evaluate pre-designed Scorpions® assays.

Read about Scorpions® technology.

Avoiding Cross Homology- For Specific Oligo Design

  • Beacon Designer™ interprets the BLAST search results and avoids the regions that have significant homologies with the database sequence.

  • Displays the homologous regions of the sequence graphically in the Sequence View pane to help visualize where the primers and probes lie with respect to them.

  • Oligos binding to regions of the sequence other that the target leads to mispriming and false positive signals.

  • It is recommended that you run a BLAST search previous to performing a primer-probe search to increase their efficiency.

  • You can choose the genome category against which you want to BLAST search, E value of the matches (by default set to 10), whether or not you want to apply filters to the query sequence (to mask repeats and low complexity regions) and the format of the output report.

  • Homologous regions to be avoided while designing oligos can be selected on the basis of either the identity of the reported hits or the e value or both.

  • You can BLAST search the primers/probes or the amplicon against a genome to verify the specificity of the design.

  • For researchers working with confidential data or who do not have a fast internet connection,
    Beacon Designer™ can interpret the BLAST results from a local database.

  • You can install a local custom database using the WWW Standalone BLAST facility of the NCBI on a server machine with Unix/Mac OS X.

  • You can also set up a local database of sequences/genome on your own machine. This option is present as "Desktop BLAST". Desktop BLAST is easy to configure and use.

Avoid Template Structures- Increase Primer Efficiency

  • Primer extension may be hindered due to the presence of template structures at extension temperature.

  • To avoid all such regions, where the template may fold upon itself, Beacon Designer™ does a template structure search.

  • You can also specify the temperature at which you would like to fold the template.

  • The results are automatically interpreted and the regions are avoided while designing primers.

  • Template structures are displayed graphically in the Sequence View pane to aid visual inspection. The bases involved are underlined in orange.

Oligo Rating

  • Beacon Designer™ rates all the primers and probes that it designs based upon how closely they match the target values specified for primer and probe search.

  • You could specify a range of tolerance for each parameter. Beacon Designer™ does not report oligos that fall outside this criteria.

  • When Beacon Designer™ finds an oligo with all parameters at the exact specified values, it has a 100 rating. If all the parameters are at their tolerance limits, the oligo rating is 0.

  • Rating is a quality indicator and is subjective in its application. It however does not means that an oligo with a higher rating will always outperform an oligo with a lower rating or that one should never choose an oligo that has a poor rating.

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