In the past several decades, in the biological engineering industry, we use the engineering microbe to produce what we expect. To meet our diverse needs, the microbe should be transformed with various exogenous genes, and the most popular vector for this job is the plasmid. There are many advantages of the plasmid vector, such as the convenience and its high transforming efficiency, but there are still many problems in the process of using the plasmid vector, one of which is the plasmid-loss, the most common and serious problem. Imagine that we transform a vital gene into the microbe in our biological lab, only to find that the plasmid with the gene get lost after generations. Things like that always upset and trouble the scientists, not to mention the industrial factories. [State the problem]

To figure out the possible reasons of the loss and find out a feasible solution, the mechanism should be understood. According to the research, the plasmid stability can be divided into plasmid structural stability and the plasmid segregational stability [1]. The former is because of the genetic mutation in the plasmid and the latter is because of random distribution of the plasmid during the cell division. Once the plasmid is unstable, there will goes the problem called plasmid-loss. Actually, in the industrial fermentation processes, the problem of plasmid segregational stability is more common, so our project focuses on the segregational instability phenomenon. After an unequal distribution, the cells divide into two groups, one with high plasmid concentration and the other with low concentration. Since the high plasmid concentration in microbe leads to low growth effect and high production rate while low concentration will lead to high growth effect and low production rate, the low concentration ones consume more but produce less. We label these black sheep as ‘slackers’. [How the problem goes]

For many years, researchers come up with various ideas to prevent the birth of these ‘slackers’. The most widely used way is antibiotics control, which will kill the ‘slacker’ whose plasmid doesn’t integrate the antibiotics resistance gene. Thus, adding antibiotics will kill the ones without enough plasmids. But this way complicates the industry procedure and results in high cost. There is also a selfish plasmid system utilizing the toxin-antitoxin system to kill the ‘slackers’ but limited for low plasmid concentration microbe. [Previous work to prevent the plasmid-loss]

Based on these background, we decide to solve these problems by employing the synthetic biological genetic circuit. Because it is more accurate, controllable, and intelligent. [Introduction to our project]