problem with overexpression

1 post / 0 new
gdbiomed's picture
problem with overexpression

I could like to introduce myself as gokila doing  doctorate  in proteomics lab . just  a few months back ,i have joined the lab and my starting  work is to  overexpress  the  recombinant protein  which is in BL-21 strain. This overexpression was  already done by two people ,now  i have been contuining it.., THE FIRST PERSON  CLONED AND OVEREXPRESSED THE PROTEIN AT CONC. 60 MICROMOLAR IPTG .THE SECOND PERSON CARRIED OUT THE WORK AFTER 1 YEAR - OVEREXPRESSED AND PURIFIED THE PROTEIN AT CONC. 0.1 MILLIMOLAR IPTG . I HAVE TRYED BOTH THE CONDITIONS WHAT THEY HAVE MENTIONED IN THEIN REPORTS  - 3 HOURS GROWTHING HTE CELLS AND INDUCED IN THE OD 0.7 TO 0.8 AND INDUCED   LEFT FOR 4 HOURS AND ANALYSED. BUT ,I  AM NOT GETTING THE INDUCED BAND AT 62KDA  . 

                                       Please help  me to get start with my doctorate work ..,