Re: [Synthetic Biology] Next-generation DNA synthesis references?

[Bryan Bishop]

On Wed, Dec 15, 2010 at 2:28 PM, Cathal Garvey wrote:
> That would be this technique, then?
> http://pubs.acs.org/doi/abs/10.1021/ja030376n

Thank you. There are also a few others on that related path:

Solid-Phase Oligodeoxynucleotide Synthesis: A Two-Step Cycle Using Peroxy Anion Deprotection
http://pubs.acs.org/doi/abs/10.1021/ja030376n
abstract: "A novel solid-phase phosphoramidite based oligodeoxynucleotide two-step synthesis method has been developed. Keys to this method are replacement of the 5‘-dimethoxytrityl blocking group with an aryloxycarbonyl and the use of N-dimethoxytrityl protection for the exocyclic amines of adenine and cytosine. With these modifications, coupling of each 2‘-deoxynucleoside 3‘-phosphoramidite to the growing oligodeoxynucleotide on the solid support can be followed by treatment with an aqueous mixture of peroxy anions buffered at pH 9.6. This reagent effectively removes the carbonate protecting group and simultaneously oxidizes the phosphite internucleotide linkage. As a consequence a new two-step synthesis cycle is possible. Oligodeoxynucleotides synthesized using this approach are identical to authentic samples when tested by a variety of analytical techniques."

Synthesis of pentathymidylate using a 4-monomethoxytritylthio (MMTrS) group as a 5′-hydroxyl protecting group: toward oligonucleotide synthesis without acid treatment
http://dx.doi.org/10.1016/S0040-4039(01)01873-1

A new protecting group for 5'-hydroxyl function of nucleotides in oligonucleotide synthesis without acid treatment utilizing unique properties of tritylthio group
http://nass.oxfordjournals.org/content/2/1/27.abstract
which says: "... Furthermore, they can reduce the reaction steps required for a chain elongation cycle from four to three because the deprotection of the 5′-hydroxyl protecting group and the oxidation of the phosphite intermediate can be performed simultaneously by a single aqueous iodine treatment."

Synthesis of oligodeoxynucleotides using the oxidatively cleavable 4-methoxytritylthio (MMTrS) group for protection of the 5′-hydroxyl group
http://pubs.rsc.org/en/Content/ArticleLanding/2010/NJ/b9nj00678h

New Protected Protecting Groups for the 5′-Hydroxy Group of Deoxynucleosides by Use of 2-(Hydroxymethyl)- and 2-[(Methylamino)methyl]benzoyl Skeletons and Oxidatively Cleavable Tritylthio and (4-Methoxytrityl)thio Groups
http://onlinelibrary.wiley.com/doi/10.1002/hlca.200490207/abstract

A new strategy for the synthesis of oligodeoxynucleotides directed towards perfect O-selective internucleotidic bond formation without base protection
http://dx.doi.org/10.1016/j.tetlet.2003.10.156

patents:
Precursors for two-step polynucleotide synthesis
Methods and compounds for polynucleotide synthesis

> I'd say we'll see the first programmable cell before we see a true,
> non-iterative improvement in chemical synthesis. The minute I have enough
> money to start prototyping, I'll make that priority #1. There has been a
> disappointing lack of improvement in the price of chemical DNA synthesis in
> the last few years. It's either down to limitations on economy-of-scale for
> conventional DNA synthesis, lack of improvement in technique, or price
> fixing. Doesn't matter which really, it looks like the field is due for a
> dramatic shift rather than an improvement on the status quo.

On an unrelated note, I am not convinced that oligo libraries + ligation is a suitable alternative to developing better DNA synthesis techniques. Maybe someone knows of a paper/argument that could convince me that oligo libraries are a better idea than improving DNA synthesis?

- Bryan
http://heybryan.org/
1 512 203 0507

[Raik Gruenberg]

Hi Bryan,

check out the technology used by Sloning Biotechnology. They do
repetitive IIS restriction / ligation on solid support and elongate
their fragments by 4 bp each cycle. Apparently this didn't translate
into cost-savings though but allows them to synthesize stuff where the
classic methods fail (e.g. high repetitions) and they have some
advantage when it comes to the synthesis of finely tuned libraries.

Another German company, Febit, synthesize dense oligo arrays within
microfluidic channels and, at some point, were also trying to market
this as a table-top DNA synthesis platform. I think that line of
business didn't take off though.

Then there is the recursive DNA synthesis from the people at Weizmann.
Really cool stuff, you should check it out. They are depending on
Oligos as input though (but can also use any longer
fragments/templates that you already have). I just run into the lead
author, Gregory Linshiz. His former lab is trying to commercialize the
technology.

High throughput sequencing technology like Ilumina is actually based
on watching the (template-driven) synthesis of single DNA molecules. I
don't remember which company the talk was from, but apparently there
are some ideas to re-use these kind of technologies for actual
synthesis.

Hope that helps!
Greetings,
Raik

[Bryan Bishop]

On Wed, Dec 15, 2010 at 5:29 PM, Raik Gruenberg wrote:

check out the technology used by Sloning Biotechnology. They do
repetitive IIS restriction / ligation on solid support and elongate
their fragments by 4 bp each cycle. Apparently this didn't translate
into cost-savings though but allows them to synthesize stuff where the
classic methods fail (e.g. high repetitions) and they have some
advantage when it comes to the synthesis of finely tuned libraries.

Karl Handelsman suggested I check out Sloning (at first I thought he made a typo). The only technical details I see are here:

http://www.sloning.com/index.php?option=com_content&task=view&id=9&Itemid=32
http://www.sloning.com/images/sloning/slonomics_large.png

"Unlike traditional methods which rely on individually designed and synthesized single stranded oligos, which have to be joined (ligated) together, Slonomics® uses a library of pre-made double stranded triplets that act as universal building blocks sufficient for thousands of gene syntheses processes. The triplet library represents all possible sequence combinations necessary to build any desired DNA molecule. Consisting of double stranded DNA triplets, these standardized building blocks, are combined by means of a series of validated reaction steps (see Figure). In the first phase of Slonomics®, the triplet building blocks are sequentially ligated in a series of cycles referred to as elongation. The output of this elongation phase is a large number of sub-fragments of the target sequence referred to as E-blocks. Together these E-blocks comprise the complete target sequence."
 

Then there is the recursive DNA synthesis from the people at Weizmann.
Really cool stuff, you should check it out. They are depending on
Oligos as input though (but can also use any longer
fragments/templates that you already have). I just run into the lead
author, Gregory Linshiz. His former lab is trying to commercialize the
technology.

Recursive construction of perfect DNA molecules from imperfect oligonucleotides
http://www.nature.com/msb/journal/v4/n1/full/msb200826.html
pdf: http://www.fractal.org/Life-Science-Technology/Publications/Recursive-DNA.pdf

De novo DNA synthesis using single molecule PCR
http://nar.oxfordjournals.org/content/36/17/e107.full

("Single molecule PCR" is music to my ears.)
 

High throughput sequencing technology like Ilumina is actually based
on watching the (template-driven) synthesis of single DNA molecules. I
don't remember which company the talk was from, but apparently there
are some ideas to re-use these kind of technologies for actual
synthesis.

That's especially what I am interested in finding out about- could you help me find anyone doing that or talking about that? Alternatively, Cathal offered a suggestion earlier today for DNA synthesis methods where parts of the process are enzymatic (like with TDTs).

thank you :-)

[Mackenzie Cowell]

Recursive construction of perfect DNA molecules from imperfect oligonucleotides
http://www.nature.com/msb/journal/v4/n1/full/msb200826.html
pdf: http://www.fractal.org/Life-Science-Technology/Publications/Recursive-DNA.pdf

Interesting paper.  The supplemental material for the "robot control language" pdf mentioned in the paper doesn't appear to be online.  Has anyone found it?  It's supposed to be at http://www.weizmann.ac.il/udi/papers/rpl.pdf

Cheers

mac

--
+1.231.313.9062 / m...@diybio.org / @100ideas

[Bryan Bishop]

On Mon, Jan 31, 2011 at 4:14 PM, Mackenzie Cowell <m...@diybio.org> wrote:

Interesting paper.  The supplemental material for the "robot control language" pdf mentioned in the paper doesn't appear to be online.  Has anyone found it?  It's supposed to be at http://www.weizmann.ac.il/udi/papers/rpl.pdf

is it this?
http://www.wisdom.weizmann.ac.il/~udi/papers/RobolabUserManualV1.pdf

--

[Mackenzie Cowell]

I think so, thanks.

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