Judging/Variance/Johns Hopkins-BAG

From 2009.igem.org

Contents

Request

Dear iGEM Team,
I am writing to you on behalf of the JHU BAG iGEM team to request a variance from the BioBrick standard of biological parts and assembly of biological parts. Our iGEM team has focused on developing new technologies and standards for synthetic genome construction. Our method for assembling genomic pieces depends on a new standard we refer to as the “Building Block”. The workflow of our system is as follows:

  1. Optimized protocols to create “Building Blocks” (BBs) by Overlap Extension/Assembly PCR from oligomers.
  2. USER (uracil specific excision reaction) protocol to join BBs into smaller chunks
  3. Homologous recombination in vivo to assemble smaller chunks to larger chunks, and using a second round of homologous recombination to assemble larger chunks into chromosome arms and full chromosomes

The purpose of our team is to develop and optimize techniques for large DNA construction as well as further the effort of the construction a synthetic Saccharomyces cerevisiae. Our team also has designed several novel computer programs for high throughput analysis of trace files and other sequence information. We fully understand the uniqueness of our team and we will be emphasizing that our central goals are somewhat distinct from those of most iGEM Teams (we are building a complex chassis, not stringing together parts into devices). All our methods will be described on our wiki from this perspective.

We hope that our team can provide new tools, both wet and dry, to broaden the perspective of iGEM and provide a generic toolset for the synthetic Biology community.

Advantages of our Building Block Standard:

  • Unlike BioBricks - No scars; seamless construction. Scars are not consistent with our genome redesign strategy because our knowledge of the rules of genome structure and function is insufficient.
  • DNA sequence requirements at joints are extremely flexible and can be found in any sequence; consensus of joining sites is A {Nx} T where x= 3,5,7,9 or 11
  • Much larger DNA pieces such as whole chromosomes may be assembled, with no constraints on restriction enzyme site positions.
  • Building blocks that are assembled can more easily contain more parts (gene, promoter, rbs, etc.), and systems of parts of very different sizes


Disadvantages:

  • Lack of standard Biobrick assembly standard


Difference from BioBricks:

  • Building blocks used for genome assembly, may contain more than one part or may containing partial pieces of parts. This can be viewed as an advantage or a disadvantage, depending on perspective.


Respectfully submitted,
James DiCarlo,
JHU - BAG Team

Judges' Response

Hi James,

Thanks for your email.

Quick question.

Please clarify if you are only requesting a variance from one of the ~6 existing BioBrick physical assembly standards, and if so please confirm that what you are proposing is different.

You can find the existing BioBrick technical standards online here: http://openwetware.org/wiki/The_BioBricks_Foundation:RFC

BBF RFC #29 may be a good place to start.

Alternatively, is your team proposing a new physical assembly standard as part of its iGEM 2009 entry? For example, such a contribution would satisfy option 3 of the iGEM gold medal requirements. See: http://2009.igem.org/Judging/Judging_Criteria

If you are proposing a new physical assembly standard then you should format your new assembly standard following the standard BBF RFC template. Please see BBF RFC #0 in this case.

All best! Drew

Team Response

Hey Dr. Endy,

Sorry for my delay. We plan on proposing a new physical assembly standard. With this standard we would additionally like to submit the genomic parts we have created with this method. Since we plan to create a new RFC, we were wondering about the specifics on the due dates and requirements of this procedure.

Thanks again,

James

Judges' Response

James,

Great.

When do you estimate that you could complete an RFC?

Requests for Variance are due 18 September.

DNA Submission is due 21 October.

Presumably, we could note the receipt of your request in advance of the 18 Sept date, but would need to have everything in place before the 21 October date.

The risk that you would assume here is that if something went wrong on your end with the new approach (either with the approach itself or with its documentation via an RFC) then you wouldn't have time to revert.

Best, Drew

Team Response

Hey Dr. Endy,

We could probably have a complete RFC done by the 18th, but I forgot to ask a few things in my previous email. The Building Block standard we will propose will not allow full interchangeability of parts in the same way as current standards. However it will allow interchangeability of say all promoters, all RBSs, all coding sequences, etc in a Building Block with all other items in a another Building Block as a fixed pieces. Would the limit on interchangeability automatically nix our shot at an RFC?

Thanks again

James


Judges' Response

James,

You'll be able to get a RFC.

One point of the RFC process is to get things out in the open so that people can learn from each other and figure out what works best. Another point is to have a published record of all proposed standards so that tool builders and others can refer to something specific.

Cheers / best, Drew