Is there a User Manual for Primer Express® software?

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No. However, there is a Primer Express® User Manual and an Applications Tutorial that gets loaded into the Primer Express® software folder when the software is installed onto your computer. In addition, there is the Getting Started Guide: Primer Express® Software Version 3.0 and tutorials available on the Primer Express product page.

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In the Primer Express® software there is a category called the Penalty Score. What is the definition of the Penalty Score, and should I take it into consideration when picking primers and probes?

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When creating primer/probe sets, the Primer Express® software will assign each set a Penalty Score. This score is based on the percentage of the preferred parameters, as defined on the Parameters page, that are met by each design. However, since the TaqMan®-specific Primer/Probe Document only chooses primer/probe sets that meet the Parameters criteria, the penalty score is irrelevant in that case. The score does not and will not determine the primer/probe performance. When following the default parameters set up in the software, any of the primer and probes selected by the software should perform well when utilizing the universal cycling conditions and the TaqMan® Universal PCR Master Mix.

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231141b34c82aa95e48810a9d1b33a79_FAQ

What is a primer-dimer? Can I predict possible primer-dimers for a given primer set in the Primer Express® software?

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Primer-dimers, usually undesirable, result when one primer binds to the other primer and possibly gets extended. The main disadvantages of this is that there will be background sequence or signal in your PCR or Real-Time PCR results, and the primers become unavailable for binding to the true desired target sequence. To view potential primer-dimers from a selected primer set, go to "Options", and "Show Primer Secondary Structure". The Primer Express® software has built in features to exclude extensive primer dimer formation.

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How can I convert my sequence from the positive strand to the negative strand in Primer Express® software?

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Highlight the entire sequence. On the menu bar, go to Edit --> Copy Complement and paste it into a new sequence window.

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How can I create an additional Archive file in the Primer Express® software?

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When you launch the software, hold down the "Option" key on a Macintosh computer, or hold down the "Alt" key on a computer running a Windows OS. The dialog box will lead you to create a new archive file.

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How do I save the primers and probes listed in the Primers Tab search results in Primer Express® software?

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Click the Save button at the bottom of the Primers page. Open the saved tab delimited text file from within a spreadsheet program, such as Microsoft® Excel® software. You may view and modify the file here.

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Within the Primer Express® software, what is a "GC Clamp" and what is its impact if I have a GC Clamp at the end of a primer?

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"GC Clamp" is the number of G or C residues at the 3' end of a primer. In the Primer Express® software, the GC Clamp can be set to any value, including zero (which is the default). In some cases, a GC Clamp of greater than three will cause no primers to be chosen. When designing primers and probes for Real-Time PCR it is recommended not to have a GC clamp on the 3' end of the primers.

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Where can I find a selected amplicon melting temperature in Primer Express software?

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You can find the amplicon in the Amplicon Tm column in the Primers and Probes Tab window.

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ac5dab2e99eee9cf9ec672e383691302_FAQ

When manually selecting primer and probe sequences using the Primer Express® software, is there a feature that can evaluate their compliance with Real-Time PCR primer and probe design guidelines?

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Yes, there is a feature to evaulate complaince of your primer and probe sequences. By copying and pasting the potential primer or TaqMan® TAMRA probe sequence into the Primer Test Document, you can get % G + C and melting temperature (Tm) information. The Primer/Probe Test can be found by going to TOOLS > PRIMER PROBE TEST TOOL.

If evaluating an MGB probe, you must use the TaqMan® MGB Quantification option. Guidelines for primer and probe design must be followed as outlined in the Primer Express® software v3.0 User Manual, which can be found in the Primer Express® software folder on your computer. You can find a copy on our website as well.

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I get the following message when trying to start the Primer Express® software: “Error opening Archive file: Primer Express is unable to open the archive file. Please move this file to another directory and run the Primer Express® software again to create a new Archive.” How do I solve this problem?

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This error message can occur while attempting to install the Primer Express software, or sometimes when launching the software after it has been installed on the computer. To create a new archive file, hold down Alt key (to the left of the space bar) while opening the Primer Express® application. You should get the browser window shown below. Click on “New” to create a New Archive.

The name “PXArchive” will automatically populate in the file name field. You may keep that name or change it to something different. Once you've named the Archive file, click on save. You should now be able to continue with the installation or use of the Primer Express Software.

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ee26fc66b1369c7625333bedafbfcaf6_FAQ

I am trying to install Primer Express® Software, version 2.0 on my new computer with Windows® XP as the operating system. It is giving me an error message, "Could not open Primers Express Preferences, because there is not enough memory" when trying to install or open the software despite this being a new computer. What can I do?

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The Primer Express® 2.0 software was developed to work on Windows 2000 operating systems. The "not enough memory" error message is generated due to incompatibility with the newer processor in the computers and the algorithm of the Primer Express® v2.0 software. We have developed a patch to help over come this error message. You can download this patch from our website by going to the software downloads option under the Technical Resources page on our website. Click on ABI PRISM® 7000 Sequence Detection System, then click on the PC link next to Primer Express® P4 Enabler. Also download the installation instructions by clicking on the Read Me link.

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