Team:Fudan/Parts
In order to fulfill our experiments and testing our device, we built dozens of biobricks. We devoted a lot to these vectors, and these vector works well in our project. However, our project is such a pioneer work that we have to explore different approaches to solve our problem. As a result, our vectors may combine diversified origin, which makes them really hard to be modified to the standard RFC. Though we works really hard these week, we cannot finish all the site-mutation work, especially those big long parts. Till now, we have modified half of our parts to the RFC10, which is still a large collection.
Here is the list of parts that works well in our projects!
All of our parts to build the “Ouroboros"are all proved to be working as expected, as the testing experiment. (For more information about our experiments, please check our results and notebook pages!) The testing of ouroboros is based on the complete part of cyclizing device（BBa_K1777016）, which is constructed based on many basic parts(BBa_K1777006, BBa_K1777001, BBa_K1777002, BBa_K1777003, BBa_K1777015.
Our mir-21 reporter also work as expected, which validate our design of mir-21 binding sites(BBa_K1777000). (see our design page)
We also use our cyclizing device to simulate the prototype in research program, in which we measured the half-life time of circRNA generated by our device(BBa_K1777015). (see our results page)
We also improved several parts, and design several parts based on the idea of other parts. For more information, please see our Description page.
We develop several linker based on former linker(BBa_K1486003). For part BBa_K1777017，We have optimized the codon for mammalian, and we insert a NLS to promote locating in cell nucleus. Besides ,we lengthen this linker to provide better separation of two domains. At last we avoid tandem repeat sequence(like GGGSPKKKRKVGGGS), which will facilitate fusing this linker to your domains via overlap PCR. (Part BBa_K1777005, BBa_K1777018 are also designed based on similar consideration)
Apart from that, we use beta-globin intron to provide SA and SD sequence in our circular RNA device(BBa_K1777001, BBa_K1777002, BBa_K1777003), which is inspired by the beta-globin intron parts（BBa_K404119）. Beta-globin intron provides long flanking intron for circularizing exon and can increase the expression level. We use this character of Beta-globin intron to improve our circRNA expression level.
Apart from all the parts listed above, we have acRNA biobricks and a dozen of cyclizing protein biobrick.( BBa_K1777008, BBa_K1777009, BBa_K1777010, BBa_K1777011, BBa_K1777012, BBa_K1777013, BBa_K1777014, BBa_K1777004, BBa_K1777005, BBa_K1777017 )