Hi all!
I need help!!
Actually I am preparing a cDNA library for 454 sequencing and I am using the library prep protocol and reagents from Roche. The library input looks really fine (on my eyes...I am not very expert), but the outpout very bad! It is like after the preparation I have really big concatamers that I suppose are cDNA amplicons of my library that are ligating each other! But why? I exactly followed their protocol except for the cDNA synthesis cause we have our home-made established protocol. In the first slide you can see the input libraries (are different preparations, I used the one in lane 4) and in the second how it looks after the Rapid Library Prep protocol.
If someone had the same problem, please let me know. From Roche they say is my library.
Thanks to all!
I need help!!
Actually I am preparing a cDNA library for 454 sequencing and I am using the library prep protocol and reagents from Roche. The library input looks really fine (on my eyes...I am not very expert), but the outpout very bad! It is like after the preparation I have really big concatamers that I suppose are cDNA amplicons of my library that are ligating each other! But why? I exactly followed their protocol except for the cDNA synthesis cause we have our home-made established protocol. In the first slide you can see the input libraries (are different preparations, I used the one in lane 4) and in the second how it looks after the Rapid Library Prep protocol. If someone had the same problem, please let me know. From Roche they say is my library.
Thanks to all!