Hi all....we're using the pDONR222 vector for sequencing cDNA inserts from libraries generated from the Invitrogen CloneMiner cDNA construction kit. Generally we've had no problems with this set-up but we were talking to another 3rd party recently who suggested that it might not be the most suitable vector for sequencing. It has 2 recombination sites which share near identical homology to one another and it has been suggested that if the insert is less than 1kb then these can form secondary or hairpin structures, essentially denying the primers template to prime off. There is supposed to be a Biotechniques article about it somewhere but I haven't been able to locate it.
Has anyone any similar experiences using this or a derivative vector?
Dr. Ciaran Fulton
Sequencing R&D Group
Almac Diagnostics Ltd
19 Seagoe Industrial Estate
Tel ('): +44 (0)28 38 39 57 22
Fax (7): +44 (0)28 38 39 86 76
Email (*): ciaran.fulton at almac-diagnostics.com
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