For routine yeast culture, the transgenic yeast strain was inoculated in 2 mL of synthetic complete (SC) medium lacking leucine (SC-Leu) containing 2% (w/v) glucose. The inoculum was cultured overnight at 30°C and 250 rpm. The culture was subsequently diluted 100-fold to an OD of 0.05 in SC-Leu supplemented with 2% (w/v) glucose and cultured for 16 h. Yeast was then harvested and sub-cultured for 24 h in SC-Leu containing 2% (w/v) galactose to induce recombinant protein production. Yeast cells were harvested by centrifugation and lysed in TES-B (0.6 M sorbitol in TE) using a Constant Systems cell disruptor at 35 kpsi and subsequently centrifuged at 11,000 g for 10 min at 4°C. The supernatant was then transferred to a new tube and centrifuged at 125,000 g for 90 min at 4°C. Finally, the pellet containing microsomes was resuspended with TEG buffer (20% (v/v) glycerol in TE). Recombinant enzymes were detected by immunoblot analysis using α-FLAG M2 (Genscript) detected with SuperSignal West Pico Chemiluminescent Substrate (ThermoFisher Scientific) (Supplementary Fig. 9 ).
α flag m2
Manufactured by Thermo Fisher Scientific
The α-FLAG M2 is a highly specific monoclonal antibody that recognizes the FLAG epitope tag. It is commonly used for the detection, purification, and immunoprecipitation of recombinant proteins expressed with the FLAG tag.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using α flag m2
1
Yeast Recombinant Protein Production
2
Yeast Recombinant Protein Production
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For routine yeast culture, the transgenic yeast strain was inoculated in 2 mL of synthetic complete (SC) medium lacking leucine (SC-Leu) containing 2% (w/v) glucose. The inoculum was cultured overnight at 30°C and 250 rpm. The culture was subsequently diluted 100-fold to an OD of 0.05 in SC-Leu supplemented with 2% (w/v) glucose and cultured for 16 h. Yeast was then harvested and sub-cultured for 24 h in SC-Leu containing 2% (w/v) galactose to induce recombinant protein production. Yeast cells were harvested by centrifugation and lysed in TES-B (0.6 M sorbitol in TE) using a Constant Systems cell disruptor at 35 kpsi and subsequently centrifuged at 11,000 g for 10 min at 4°C. The supernatant was then transferred to a new tube and centrifuged at 125,000 g for 90 min at 4°C. Finally, the pellet containing microsomes was resuspended with TEG buffer (20% (v/v) glycerol in TE). Recombinant enzymes were detected by immunoblot analysis using α-FLAG M2 (Genscript) detected with SuperSignal West Pico Chemiluminescent Substrate (ThermoFisher Scientific) (Supplementary Fig. 9 ).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!