In 2019, after completing my master's degree at the Institute of Microbiology, CAS, I joined Professor Sheng Yang's lab at the CAS Center for Excellence in Molecular Plant Sciences to pursue my Ph.D. When we discussed my research project, Professor Yang said with a hint of mystery, “I have the world’s most powerful cellulose-ethanol yeast strain. Would you like to figure out why it’s so robust and high fermentation performance?” Eager for a challenging project, I enthusiastically agreed, fueled by excitement and optimism. Little did I know just how challenging it would prove to be, even making me consider giving up halfway.

 The star of our story, the world’s most powerful cellulose-ethanol yeast, is called “Strawbrew”. It’s a high-performance strain, customized through selective adaptation, that has already been industrial production. You can read more about Strawbrew's origin in our paper or Professor Yang’s blog (Engineered yeast good at straw-sugar conversion). When I took on the project, senior lab members had already identified three beneficial target genes, integrating them into a clean background strain to create three engineered strains. But tests showed their fermentation performance in synthetic media with sodium salt inhibitors was far from matching that of Strawbrew. My mission was to find the remaining key targets and reconstruct Strawbrew’s superior traits, unraveling the genetic basis behind its remarkable stress resistance and fermentation efficiency.

 At that time, Strawbrew’s 288 mutations left us without a clear direction. Dr. Y.M. Liu in the lab evolved the strains with three beneficial targets using the same approach applied to Strawbrew, hoping to identify shared mutations through genomic sequencing analysis—our only clue. The Strawbrew project, which began in 2010, was nearly a decade old by 2019, and I was determined to finish this “relay race” with a perfect finish.

 Though the experimental process was challenging, it proceeded relatively smoothly. When I added three new mutations one by one to the engineered strains, they began to perform comparably to Strawbrew in synthetic media with sodium salt inhibitors! I was ecstatic and immediately emailed Professor Yang and other project advisors to share the news. But, I had to admit, the strain wasn’t yet reconstruct Strawbrew’s phenotype in hydrolysates. I combed through literature and carefully reviewed each experiment to ensure I had missed nothing. When I discovered one more beneficial mutation from other six mutation and added it, the engineered strain closely matched Strawbrew’s performance in hydrolysates. Only by magnifying the fermentation curve could you detect any difference. Feeling discouraged, I asked Professor Yang, “Do we really need to reconstruct Strawbrew’s phenotype exactly?” He unhesitatingly answered “Yes”, encouraged me to keep pushing forward, and discussed other potential targets with me.

 Reinvigorated, I went back to verify every possible target. Finally, after supplementing the strain with multiple copies of xylose isomerase, the engineered strain fully matched Strawbrew’s performance in hydrolysates—and, in a few independent experiments, even surpassed it. This time, I felt less of the initial excitement and more a sense of calm composure.

 As I worked to decode Strawbrew, I grew into a capable doctoral researcher. Beyond learning specialized knowledge and techniques, the most valuable lesson I gained was the ability to calmly analyze disappointing results and optimistically see them as stepping stones, “Look, one more wrong path eliminated; we’re getting closer to the right one!”

 I hope my story encourages fellow students on their academic journey to stay true to their original pursuit of truth.

*The image was created using ChatGPT.*