pHBM146

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	The recombinant plasmid pHBM146 was digested with HpaI to remove the origin of replication (ori) of E. coli and the bla gene, and then was transformed into the S. cerevisiae INV strain, according to the manufactures instructions. The S. cerevisiae transformants were screened on synthetic complete (SC-Ura) medium containing 0.5% Remazol Brilliant Blue (RBB)-xylan with the replacement of glucose by D-galactose for 12 h at 28 °C.

Protocol tips

The recombinant plasmid pHBM146 was digested with HpaI to remove the origin of replication (ori) of E. coli and the bla gene, and then was transformed into the S. cerevisiae INV strain, according to the manufactures instructions. The S. cerevisiae transformants were screened on synthetic complete (SC-Ura) medium containing 0.5% Remazol Brilliant Blue (RBB)-xylan with the replacement of glucose by D-galactose for 12 h at 28 °C.

Publication protocol

The recombinant plasmid pHBM146 was digested with HpaI to remove the origin of replication (ori) of E. coli and the bla gene, and then was transformed into the S. cerevisiae INV strain, according to the manufactures instructions. The S. cerevisiae transformants were screened on synthetic complete (SC-Ura) medium containing 0.5% Remazol Brilliant Blue (RBB)-xylan with the replacement of glucose by D-galactose for 12 h at 28 °C. The transformant with the largest halo was selected for liquid fermentation. The two-stage liquid fermentation was carried out with two different media. During the enrichment stage for strains, 50 ml of yeast extract peptone dextrose (YPD) and SC-Ura mediums were used. The enrichment was carried out in a shaking incubator (250 rpm) at 28 °C. During the induction stage, the cells were harvested until the OD600 of YPD medium and SC−Ura medium reached 25 and 5.0, respectively and then transferred into 25 ml induction medium. The induction mediums were YPD medium and SC−Ura medium in which glucose were replaced by D-galactose. The initial concentration of D-galactose in the induction mediums were 2% and the induced cultivation were carried out for 4 days. Every 24 h, galactose was added at a final concentration of 2% to induce xylanase expression.

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