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| Proteins, Expression, Isolation and Analysis | The?recombinant protein?produced in?E. coliBL21(DE3) was purified using Ni-Charged MagBeads according to the manufacturer's instructions (GenScript, China). The culture of?E. coli?BL21(DE3) was concentrated to 50?ml and later centrifuged at 12,000?rpm for 1?min at 4°C. The cell pellets were re-suspended in 5?ml of binding buffer (0.1?M Tris–HCl, pH?8.0), added to 50?μl?lysozyme?(final concentration of 100?μg/ml), and incubated on ice for another 30?min. Cells were disrupted by sonication (20% of power treatment for 30?s at 4°C) and centrifuged at 12,000?rpm for 15?min at 4°C. The?supernatant?and cell pellets were, respectively, collected and detected using 12% SDS-PAGE to identify the expression of StMsn2 in?E. coli. | Get A Quote |
Background In Saccharomyces cerevisiae, Msn2, which acts as a key transcription factor downstream the MAPK-HOG cascade pathway, also regulates the expression of genes related to stress responses. However, little is known about the regulation mechanisms of the transcription factor in Setosphaeria turcica. Results In this study, a zinc finger DNA-binding protein, designated as StMSN2, was cloned from S. turcica. Sequencing results showed that StMSN2 had a 1752 bp open reading frame (ORF), which was interrupted by an intron (135 bp) and encoded a putative 538-amino acid protein. Phylogenetic analysis further revealed that StMsn2 was more closely related to Msn2 of Aspergillus parasiticus. StMSN2 was cloned into t... More
