Repeat Expansion Diagnostic Techniques Part 2 - Repeat Primed PCR

This is Part 2 of a 2-part series about molecular diagnostic testing for repeat expansion disorders. In this video, triplet-primed PCR is explained using Fragile-X disorder (FMR1) as an illustrative example.

Repeat Expansion Diagnostic Testing: Part 2 - Triplet-Primed PCR by Navneet Aujla and Talia Silver is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.

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[Proteinace]

The reverse primer is FAM-labeled, not the forward. If the flanking gene-specific forward primer were FAM-labeled, you wouldn't see the repeat-primed amplicons on CE. Finally, the repeat primer does not overlap the flanking sequence. It does in some assays (this is the basis for "anchor" priming), but not here.

[HoiLamDoong]

These videos are very clear and helpful. Hope more videos like this can be made! Thank u very much.

[ari.sandoval]

Thanks for the video! it was very useful for me, i work with myotonic dystrophy type 1 in Mexico :D

[aadilyousufdar]

Thank you for the video! But I could not understand how forward and reverse (flanking) primers are able to amplify PM and FM allele while in conventional PCR the same two primers do not.

[jongelb]

So if I'm getting this right the forward and reverse primers are intended to copy the entire length of the gene. The forward primer binds to the template/antisense strand, while the reverse binds to the coding/sense strand. Finally the repeat-specific primer randomly generates repeats of variable lengths.

[windflow]

How to design a Repeat Primed PCR for repeat expansion disease? Can you add a video tutorial of such primer design ？