TY  - JOUR
TI  - Expression and Purification of the Eukaryotic MBP-MOS1 Transposase from Sf21 Insect Cells
AU  - Jaillet, Jérôme 
AU  - Dussaussois-Montagne, Audrey 
AU  - Renault, Sylvaine 
AU  - Augé-Gouillou, Corinne 
VL  - 4
IS  - 20
PY  - 2014
DA  - 2014/10/20
SP  - e1262
C1  - Bio-protocol 2014;4:e1262
DO  - 10.21769/BioProtoc.1262
UR  - https://doi.org/10.21769/BioProtoc.1262
AB  - Here, we present the full-length protocol for purifying the recombinant MOS1 transposase from insect cells used in our recent publication (Pflieger et al., 2014), which involved a N-terminal MBP-tag and maltose-affinity chromatography. Due to their overall basic properties, transposases are often difficult to purify, especially because they tend to aggregate. Since the 90s, we chose a method of purification without a denaturation step. Our first priority was to preserve the 3D structure of the protein in order to maintain its biochemical activities with the highest specific activity. Nevertheless, our production/purification made from bacteria regularly contain truncated products (or degradation products) and their levels increase with concentration of purified transposase. In contrast, production/purification made from eukaryotic cells do not contain such degradation product. We thus developed a protocol involving the pVL1392 baculovirus transfer vector and the BaculoGoldTM baculovirus expression system, allowing the expression of recombinant MOS1 from baculovirus-infected Sf21 cells.
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False