Protein Expression in E. coli using pEX Expression Vectors

1. Protein Expression in E. coli using pEX Expression Vectors • Validation Data

2. 5 New E. coli Expression Vectors Please see vector maps at http://www.origene.com/cdna/TrueORF/DestinationVector.aspx then click on “Bacterial expression” tab

3. Protein Expression in E. coli N-tagged vector 4 different mammalian ORFs was cloned into the pEX-N-His-GST vector. GST-fused proteins were expressed at high levels with all constructs.

4. Protein Expression in pEX-C-His Vector Protein Expression in pEX-C-His Vector 1 2 MW (kDa) 191 97 The ORF of Bin1 was cloned into pEX-C-His vector and expressed in BL21(DE3) cells. Total protein lysates were loaded on the SDS-PAGE gel. 64 BIN1 51 39 28 • pEX-C-His-Bin1 (NM_139351) no IPTG • pEX-C-His-Bin1 (NM_139351) with IPTG

5. Highly Efficient Protein Expression pEX-N-His-GST vector shows comparabe high expression level of GST with pGEX-2T vector.

6. The Fusion Tag Can Be Cleaved A purified GST fusion protein was incubated with TEV protease at 30oC for the indicated period of time. The recombinant protein and GST tag were separated by TEV cleavage (C).

7. pEX-1 for Untagged Expression When no-tag is desired for the recombinant protein, pEX-1 is the vector of choice, appending only two amino acid (T and R) to the C-terminus of the protein.

8. Easy TrueORF Shuttling into pEX 37,000 TrueORF clones can be easily shuttled from entry vector into E. coli pEX vectors. The entry and destination vectors are digested with two rare cutters: Sgf I and Mlu I or other specified enzymes. For pEX-1 and pEX-C-His vectors use BseR I and MluI for subcloning. *

9. Order TrueORF in E. coli Vectors • For all destination vectors, there are two options: • Purchase both the TrueORF in pCMV6-Entry vector and the desired destination vector and follow the simple shuttling protocol. • Ask OriGene experts to perform the shuttling. OriGene will provide highly purified ready-to-use plasmids.