When constructing a controllable inducing lysis system in Synechocystis sp. PCC 6803, we need the P22 genes 13、19 and 15.，But to synthesis them is not an easy work. Fortunately, we got access to Prof. Liu from East China University of Science and Technology, who used a plasmid containing those genes according to her paper “A controllable bacterial lysis system to enhance biological safety of live attenuated Vibrio anguillarum vaccine”. Interested in our project, she would like to give the plasmid to us. But it’s a great pity that the strain and the plasmid were lost because of the removal. Though failed to get the genes in this way, we are grateful for Prof. Liu’s help.

Dear professor, I’m Jiawei Chen, a junior student in the Academy of Chemical Engineering, Tianjin University, majoring in pharmaceutical engineering.
Recently, our team were conducting an experiment aiming to build a novel in vivo inducible bacterial lysis system in Synechocystis. We have found your journal article, A controllable bacterial lysis system to enhance biological safety of live attenuated Vibrio angulliarum vaccine, published in Fish & Shellfish Immunology last year. And we have got great interest in the regulatory switch to control the expression of phage P22 lysis cassette 13-19-15.But we don’t have a deep insight in this field of research and have several questions.
From the article we could know that these 3 genes are from Phage P22. While searching in the GenBank, however, we have found there were different kinds of Phage P22. And different phages have different genes. Thus, we want to exactly know which kind of phage these 3 genes come from. And could you please tell us the gene sequence if possible so that we can make a total synthesis of these genes?
Thank you and we hope to get your guidance.
Best regards
Jiawei Chen

Dear Jiawei Chen, These genes were brought from America and I can give you these genes. So you don’t have to synthesize them. Please let me know your address.
Professsor Liu

Hello professor, It’s excited to receive your email. Thanks a lot for your support. Not only has it solved our problems, but also saved our funds.
It is difficult for us undergraduates to conduct a project by ourselves. In the process of experiments, we will inevitably encounter a great number of problems. So your help means a lot to us. Thank you again and wish our project will go on well with the plasmid you provide.
Besides, our plan is to control gene 13 by a nickle-inducible promoter, meanwhile, make a normal expression of gene 15 and 19. In doing so, we can build a novel in vivo inducible bacterial lysis system in Synechocystis. So could you please give us the plasmid profile or relative gene sequences so that we can make further design to fulfill our purpose.
Best wishes
Jiawei Chen

Jiawei Chen,
The 13-19-15 gene sequence is in the appendix.
This lysis gene is bond with a regulatory promoter by overlap PCR and then cloned to pUTat plasmid.
Plasmid pUTat is based on plasmid pUC18, modified according to the requirement of my lab. First, we cut off lac regulatory sequence and multiple cloning site (MCS) in pUC18 using Ndel and pvull, eliminate its influence on the expression of downstream inserted fragment. Second, we insert MCS and a transcription terminator rmBT0T1 from plasmid pBR322, which is an intrinsic terminator, so that we can effectively stop the transcription of upstream gene and promote its transcription efficacy. At the same time, we change plasmid replicator into a low-copy replicator. The modified plasmid is called pUTat.
What you need to do is to design primers according to the 13-19-15 gene sequence, amplify the target gene by PCR and make it cloned into your plasmid vector in your experiment.
Professor Liu

Dear teacher,
It’s glad to get your help.
Actually, we are participating in IGEM competition, held by MIT. This year our team’s instructor is Professor Yuan, who is in great favor of our plan.
But our biggest problem is there’s not much time left for us. The summer vacation is almost over during which we pay a lot of attention to experiments. But the period of cultivation of Synechocystis is very long. And we hope we can make a meticulous plan and build a perfect model as soon as possible.
So I want to know when the administrator of strain bank will work. And could you give us your contact information if possible? We’d like to get the plasmid profile and design primers soon.
Thanks a lot for your help.
Jiawei Chen

Jiawei Chen, Sorry to tell you the administrator of strain bank didn’t found the strain and plasmid. Maybe it has been lost during the move.
Professor Liu