Dr. Haoqian Zhang

‘Science is not about the application, but about interesting.’ — Dr. Zhang

He confirmed us that the question is not whether it could be applied or not, but is it interesting, and when to get started. Boosting our confidence to move on our project as a science campaign, he presented us with the idea of co-culture. With this design we could effectively give the synthesize our product on the same plate, increasing efficiency and reducing the time spent on production. Besides this, he introduced us with a recent technology: stable promoters of TALEsp. He suggested that it could be used to stabilize and enhance the production of geraniol inside E.coli. His advice was of significant impact in the shaping of our project trajectory, and brought us with the later goal of constructing a TALEsp library.

Professor Christopher Voigt

Figure.

Having decided to use TALEsp in our project, we were not sure about whether should we apply it to yeast or not. Thus on July 7th, at the 4th Synthetic Biology Youth Scholar Forum, we asked professor Christopher Voigt about his opinion on integrating TALEsp into yeast and other technical questions related. Professor Voigt hinted that since TALEsp sequences are similar to each other, an application in yeast would most possibly trigger the reassemble system of yeast and dysfunction its ability. Also, in the aspect of E.coli TALEsp, he suggested integrating the circuits to the genome to prevent failure of TALEsp when a large number of bacteria exist (e.g fermenter). When coming into the technique of characterizing TALEsp promoters, we were uncertain of which method to use: GFP or RNA-seq. He clarified that these two techniques are not of much difference, only that RNA-seq is a little more accurate, but at the same time much more expensive. We thus decided a GFP characterization method.

Dr. Qiuli Lou

To work with yeast is a great challenge to our team. Many information of Yeast remains unclear, yet the most emergence one is that we don’t know exactly the condition to carry out colony PCR. To tackle this problem we sought help from Lou, a graduate student majoring on biotechnology at Fujian Agriculture and Forestry University. From this conversation, we figured out a way of diluting free colonies in 20mM NaOH for 10min 100˚C before the normal PCR process. Breaking through this technical problem our yeast circuits construction gets to move on.

Dr. Junbiao Dai

Our design on experiment involves knock out of 4 enzymes and knock in of 3 enzymes in yeast, and we primarily decide, in the knock out experiments, to use the Hi-crisper technique developed by Junibiao Dai and his team. (further details see our design page: ) However, after several attempts, we found the technique troublesome: they did not work as demonstrated by the paper. Also, during the knock in stage, we found out that E.coli often dies when our pYES plasmid is transformed, yet yeast often will not carry this plasmid when transformed directly from Gibson product. We thus sought for a chance to interview Dr. Dai and inquired him the problem we faced. He suggested that if Hi-crisper will not work, then a traditional knock out process is recommended to be carried out. Also, for the dying E.coli plight, he hinted that the pYES plasmid with three coding sequences of the knock in enzymes might be toxic to E.coli as a whole. The circuits heavies the metabolic burden of E.coli tremendously and leading to their death. Finally, he debugged the yeast issue, saying that Gibson product has a low concentration, and presented a feasible solution of lengthening the homogenous arm to trigger the homogeneous recombination stem of yeast. We adapted his suggestion of knock out sequence one by one.

Dr. Chong Zhang

Chong Zhang is the partner of Junbiao Dai; he introduced he and Dr. Dai’s paper: MiYA, an efficient machine-learning workflow in conjunction with the YeastFab assembly strategy for combinatorial optimization of heterologous metabolic pathways in Saccharomyces cerevisiae to us. This paper introduced a possible way to enhance the production of nepetalacol in yeast, and we grew very keen on adopting this method. However, due to the lack of database and the scanty of time, this method was obsoleted.

Dr. Akihiko Kondo

We met Dr. Kondo at the 4th Synthetic Biology Youth Scholar Forum and introduced our project to him, with whom grew very interested. He suggested his point mutation plasmid to us. Due to the lack of time, we did not integrate this suggestion into our project; but we are thankful to his help and appreciate his kindness

Rescue Team

To improve our KW, we decided to construct a 3D model; but whether is this device really aiding the works of rescue teams or not, or if it's but a burden to them, remains unanswered. Thus a few weeks later we managed a thorough online interview with the leader of the rescue team of Ms. Fenghua Li, Dachui, and introduced for the first time our thoughts of this KW to the stakeholder. To our surprise, this design was much approved by Dachui and other team members, and consolidated our will of materializing the KW.

Xin Wei

To remove this liability and improve safety factors, we finalized our cat-attracting system as a bottle of extracted essential oil and a fan to spread the odor that obviously increases the safety and ethical factor.

Safety issues have always been our first consideration; since we were, then, planning to use a plate with living organisms in, the device must be constructed deliberately and safely to prevent any leak possible. One of our public engagement presented the answer to us. On July 6th, our first presentation, also being our first event facing the public, took place at X.factor. This presentation, free in the boundary of age, background, or academic level, had brought us waves of brand new ideas that all enlightens us with revolutionary designs. Yet one of them, coming from a university student majoring in psychology, remained distinctively important and profoundly impactive:

John Henry

After the presentation, we consulted a hardware designer in X.factory about his opinion on our design. John Henry rejected our initial plan of a tent and canvas, assuring us that the transportation of the whole house would be inconvenient and troublesome; plus, an unstable structure of cat house would not serve well as a permanent household. Also, he hinted that a cat house for capturing and living should be low profile, not obvious, and functional. He thereby insists wood as the building material which provides a safe environment, sustainable residential area, and operational structure. Following his idea, we thus determined the building material and started our materializing campaign. This design bestowed a greater sustainability to our product, since now we are using a long-lasting and sediment material for constructing, which enhances the safety of household provided to cats. For further design and construction, see .

Hui Huang

Seeking possible improvements, we interviewed the leader of SSAPA (Shenzhen Small Animal Protection Association), Mr. Huang. As a pioneer of protecting animals who possess 3 years of stray cats rescuing experience, Mr. Huang reminded us of a phenomenon that none of us had ever thought of:

‘Cats will fight back wildly after being captured. Sometimes hurting themselves severely.’ — Mr.Huang

This raises our alert as the safety of cats are unconsciously threatened. Again, a change is introduced to our KW, which discards its auto closed door, and replaced with a surveillance camera and remote control door. By this course, we can detect whether is cat attracted inside the house, and thereby to close the door or not. Some other improvements are made in order to enhance the safety level of our product; details could be found in both our safety form and applied design page. Such improvement lately allowed us to conduct our final investigation of the behavior of cats smoothly.

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