Anyone know what the fill-in enzyme is?

[gaster]

Does anyone know the enzyme Roche uses for the fill-in reaction in the gsflx library prep kit? Anyone know an enzyme that does the same thing?

This is to fill-in a phosphodiester bond on one strand after adaptors are added.

Thanks!

[marklar]

have a look at the methods section in this paper
i've used this protocol successfully. step 14 is the adapter fill-in step using Bst polymerase.

[gaster]

Thanks, Marklar. I will check it out.

[lionline]

Bst polymerase is used in that paper.

Can I use Large Fragment of DNA Polymerase I (Klenow Fragment) instead? It is said to have strand displacement property.

Thanks for any comments.

[gaster]

Yes, I think you can use the 3'- 5' exo- Klenow Fragment. Check out NEB's latest flyer. I am told the Bst polymerase adds dAs.

Good luck!

[lionline]

Thanks gaster!