the MYS!S Project

View As One PageView As Slideshow     The MYS!S Project: BioBrick Optimizer Abstract MYS!S is a stand alone software tool designed to assist an “iGEMer” when designing a new BioBrick part or modifying an existing one. MYS!S handles several problems that a synthetic biologist might encounter in the lab that can be solved with site directed mutagenesis. The utilities that MYS!S supports to modify DNA sequences on paper, can be “actualized” in the lab using site directed mutagenesis to change physical DNA “stored” in a backbone. MYS!S will determine the most efficient modified sequence by optimizing for the least number of rounds of site directed mutagenesis. Once the changes are finalized primers are designed. In addition to primer design, MYS!S provides lab protocols for both site directed mutagenesis and transformation into competent cells. Currently the lab protocols are supplied as a BioCoder compliant C++ file, that can then be compiled with the BioCoder library.

Operations
I. DNA Optimization First, it performs DNA codon optimization based on the target organism by removing rare tRNAs and replacing them with more prevalent tRNAs. (We noticed that many teams decided to utilize sequences from a variety of organisms such as firefly luciferase to ultimately be transformed into an experimentally “stable” organism.)   II. Processing Secondly, the program checks the coding sequence for any restriction sites that are not allowed by the format specified by the user. After checking the sequence, MYS!S removes the restriction sites while ensuring that the modified sequence is still codon optimized. In addition, it was pointed out that frequently a synthetic biologist would like to change specific amino acids in the DNA coding sequence. MYS!S decides what difference in nucleotide sequence are required to best make the amino acid change.   III. Restriction site analysis Alternatively, an iGEMer might also want to add restriction sites to a DNA sequence in order to remove a particularly useful sequence. The program can handle this by either allowing the user to specify where the restriction site should be or by choosing what restriction site is desired and how sites need to be added to the sequence.

How Mys!S Does it
The MYS!S Environment One of the main ideas behind MYS!S was to make synthetic biology more accessible to a wider “spectrum” of people. We want the program to provide an environment that enables the user to understand and explore the changes being made to the DNA in a more intuitive manner. Custimization In addition, MYS!S is designed to be highly customizable by providing the user with the ability to add and modify components such as an organism’s codon usage table and new assembly standards. The ultimate goal is to create a program that can be customized by the user for their specific work and the lab protocols they are comfortable with.

MYS!S: An Introduction

Walk through One of the main ideas behind MYS!S was to make synthetic biology more accessible to a wider “spectrum” of people. We want the program to provide an environment that enables the user to understand and explore the changes being made to the DNA in a more intuitive manner. In addition, MYS!S is designed to be highly customizable by providing the user with the ability to add and modify components such as an organism’s codon usage table and new assembly standards. The ultimate goal is to create a program that can be customized by the user for their specific work and the lab protocols they are comfortable with. To showcase the capabilities of MYS!S we would like to walk you through an analysis of a current BioBrick in the registry. For this example we are going to use part BBa_K191006 which is the protein coding sequence for LovTAP. LovTAP was used by both of our laboratory projects Etch-a-Sketch and Full Adder. One of the issues with LovTAP is that it contains restriction sites not allowed by some BioBrick assembly standards. Lets say we want to transform the LovTAP coding region into e-coli After opening MYS!S, navigate to the screen to manage components. To do this go to the MYS!S menu and click “Manage Components”.   It will open up a screen that will allow the user to manage which organisms and assembly standards the program handles. From this screen the user can add organism codon tables, modify existing ones, and delete those not needed anymore. The same functionality applies to standards. The user can specify the prefix, suffix, and the restriction sequences that the nucleotide sequence should not contain.         Now we are going to exit the manage components screen and open a new assembly standard check. You can find the assembly standard check option by going under the file menu then click “New” then click “Assembly Standard Check”.     Next, we need to enter all the information required to perform an assembly standard check. The fields that have to be filled in are the name field, the organism, the standard, and the plasmid. Obviously the user also needs a nucleotide sequence to analyze. This sequence needs to be entered in the original DNA sequence text area. Note MYS!S requires the nucleotide sequence to start with ATG and be in frame.     After these fields are completed the user can hit the go button to perform an assembly standard check. The original DNA sequence and the modified sequence are displayed top and bottom. All proposed changes in the modified sequence are colored green the corresponding nucleotides in the unmodified sequence are colored purple. The primers needed to transform the original Lovtap sequence to the assembly standard acceptable Lovtap are shown in alignment with the 5’ and 3’ ends labeled.

Plug-ins
I. Protocols In the protocol tab is a BioCoder compatible c++ file that contains the lab procedures for mutating the original Lovtap DNA into a standard safe form. The c++ file can be compiled with the BioCoder software available here. http://research.microsoft.com/en-us/um/india/projects/biocoder/   II. Rna Structure Analysis In the RNA structure tab there are images of the unmodified and modified RNA structure. We hope that this will help the user decide whether the changes are structurally advantageous. Hopefully in the future more advanced RNA structure modeling algorithms can be implemented to help the user make an informed decision.     Unfortunately, we’re talking about the capabilities of the future MYS!S v.10, for now all we have is the beta edition. So for the time being, these are the imminent improvements we would like to make for the second version of MYS!S. Better algorithms for modifying DNA. Currently, when determining how to modify DNA MYS!S does not take into account the eventual RNA structure and whether the changes will inhibit protein production. We would like to incorporate algorithms that make changes to DNA in a way that will increase the amount of protein formed by translation. On the same note, it might also be helpful for a synthetic biologist not just to increase protein production but maybe to limit it. Not just support site directed mutagenesis. Right now MYS!S for a codon optimization creates a large number of primers for a sequence of say 700bp. We’re talking about upwards of 50 primers making site directed mutagenesis realistically impossible. We would like MYS!S to support other methods of manipulating physical DNA. Better visualization methods for RNA structure: We want the user to be able to visually check whether the RNA structure is acceptable. If it is not acceptable the user should be able to manually modify the DNA sequence to improve the RNA structure. Preference for lab protocols Not all labs do things the same, MYS!S should be able to customize lab protocols to how the user’s lab gets things done.

Where can I get MYS!S?

Github MYS!S is currently available as an Eclipse download. It will very soon be available as a java application. The downloads can be obtained from our github site. https://github.com/RutgersGEARS/iGEM-Rutgers-Software MYS!S is still a work in progress. Please feel free to report any bugs or crashes that occur as issues on our github page. https://github.com/RutgersGEARS/iGEM-Rutgers-Software/issues