The general approach begins with DNA from a variety of sources. Here we used oligonucleotides synthesized from microarrays as well as from conventional sources. Then, next-generation sequencing is used to read and identify oligonucleotides
Right now the cost of synthesizing a base [using conventional technology] is about 10 cents. That’s the current street price for raw oligonucleotides. For synthesizing simple genes, it’s more like $1.30 a base. [George Church claimed a method he had in 2007] could manufacture oligonucleotides at .01 cent per base.
Prediction from 2006, costs of gene synthesis per bases pair have fallen 50-fold, halving every 32 months. At the same time, the accuracy of gene synthesis technologies has improved significantly
The construction of synthetic biological systems involving millions of nucleotides is limited by the lack of high-quality synthetic DNA. Consequently, the field requires advances in the accuracy and scale of chemical DNA synthesis and in the processing of longer DNA assembled from short fragments. Here we describe a highly parallel and miniaturized method, called megacloning, for obtaining high-quality DNA by using next-generation sequencing (NGS) technology as a preparative tool. We demonstrate our method by processing both chemically synthesized and microarray-derived DNA oligonucleotides with a robotic system for imaging and picking beads directly off of a high-throughput pyrosequencing platform. The method can reduce error rates by a factor of 500 compared to the starting oligonucleotide pool generated by microarray. We use DNA obtained by megacloning to assemble synthetic genes. In principle, millions of DNA fragments can be sequenced, characterized and sorted in a single megacloner run, enabling constructive biology up to the megabase scale.
There are many cost components to commercial gene synthesis, and only someone who has carefully looked over the books while wearing a green eyeshade is going to have a proper handle on them. But three of the big expenses are the oligos themselves, the sequencing of constructs to find the correct ones and labor. What the Febit paper does is illustrate a nice way to tackle the first two in a manner that shouldn’t require a lot of labor.
The cost of gene synthesis, currently starting at around $0.40 per base pair for very easy and short stuff (say, less than 2Kb), tends to restrict what you can use it for. [Dr. Robison, genomics expert] have a project concept right now that would be a slam dunk for gene synthesis — but not at $0.40/bp (which I thinks I couldn’t even get for the project). Whack that price by a few factors of two and the project becomes reasonable.
The oligo cost is a serious issue. Conventional oligos can be had for around $0.08 or maybe a bit less a base. However, each base in the final construct requires close to 2 bases in the oligo set. Some design strategies might get this down a bit. However, conventional columns generate far more oligo than you actually need. An approach which has been published (but not commercialized as far as I know), is to scale down the synthesis using microfluidics. This method matches better the amount synthesized and the amount you need, though the length and quality of the oligos needs refinements from what was reported in order to be truly useful. Microarrays are a means to synthesize huge numbers of oligos, but their quality also tends to be low and the quantity of each oligo species is much too small without further amplification. Amplification schemes have been worked out, but add to the processing costs of the oligos.
What Febit and company [the above paper by George Church et al] have done is take those microarray-build oligos and screen them using 454 sequencing. The beads containing the amplicons with correct oligos are then plucked out of the 454 flowcell (with 90% success of getting the right bead) and used as starting points.
The alternative route that I predict this NewCo is likely to go down. That would be to link up with an established provider in the field. Said provider, through their salespersons and sales software, could offer each customer an option — I can build your genes for $0.40 if you want them fast or hack that down to $0.10 a base if you can wait.
In summary, I think this is a clever idea which needs to be pushed forward. But, after a long gestation in the lab, it faces a very rocky future in the production world.
More details on the George Church Approach
Development of cheap, high-throughput and reliable gene synthesis methods will broadly stimulate progress in biology and biotechnology. Currently, the reliance on column-synthesized oligonucleotides as a source of DNA limits further cost reductions in gene synthesis. Oligonucleotides from DNA microchips can reduce costs by at least an order of magnitude yet efforts to scale their use have been largely unsuccessful owing to the high error rates and complexity of the oligonucleotide mixtures. Here we use high-fidelity DNA microchips, selective oligonucleotide pool amplification, optimized gene assembly protocols and enzymatic error correction to develop a method for highly parallel gene synthesis. We tested our approach by assembling 47 genes, including 42 challenging therapeutic antibody sequences, encoding a total of ~35 kilobase pairs of DNA. These assemblies were performed from a complex background containing 13,000 oligonucleotides encoding ~2.5 megabases of DNA, which is at least 50 times larger than in previously published attempts.
The new approach relies on selective enrichment of long oligonucleotides from DNA microchips, allowing researchers to synthesize and assemble many oligonucleotides simultaneously. That, in turn, provides an opportunity to scale up and streamline the gene synthesis process.