Other Species Collections
pExpress1 Library Method Overview
Express Genomics, Gaithersburg MD, constructed directionally cloned rat libraries
in the pExpress 1 vector using a proprietary method. Briefly, the method overview is as follows:
- First strand cDNA was synthesized using an oligo-dT primer containing a Not I site
and MMLV-reverse transcriptase.
- Second strand cDNA was synthesized and digested with the Not I restriction enzyme.
- The double stranded cDNA was size selected (>1.4 kb).
- The pExpress1 vector was digested with Not I and Eco RV.
- The double stranded cDNA was directionally cloned into the pExpress1 vector with:
- the 5' blunt end of the cDNA ligated to the blunt Eco RV site of the vector, thereby destroying this site
- the 3' end of the cDNA ligated to the Not I site of the vector, thus recreating the Not I site.