3 replies to this topic

[Sequencer]

Anyone who could tell me what is the best way to sequence a dna sample with tandems/repeats? I am using the tn5 transposon... so far, not so good results!

[kean_guy]

I'd prefer to clone and sequence the DNA sample with universal primers, T4, t7, M13 etc...

This is especially important if your sequencing primers have more than one annealing sites, esp. in repeats.

Cheers

[george@CASE]

I second kean_guy ! good luck!

Don't use sequencing primers that will sit on the repeats.

[methylnick]

be careful with tandem repeats as they can be unstable. I have tried to clone alpha satelite into pGEM-T easy, it's challenging but can be done and I found that recombination can occur. This could probably pose a problem for tandem repeats also. Depending on the repeats you can add things like DMSO to relax the secondary structure in your sequencing reaction.

Good luck!

Nick