Automated gene synthesis method may streamline vaccine production

Gene synthesis has become an important tool in many fields of recombinant DNA technology, including vaccine development, gene therapy and molecular engineering. The primary method for gene synthesis is oligonucleotide synthesis, a process by which scientists are able to chemically synthesize relatively short fragments of nucleic acids with defined chemical structure. This technique is extremely useful in current laboratory practice because it provides rapid and inexpensive access to custom-made oligonucleotides of the desired sequence.

Under currently known synthesis methods, about fifty different partially overlapping oligonucleotides must first be synthesized and purified in order to synthesize an approximately 2.5 Kilo-base pair (kb) nucleic acid sequence. The known procedures require at least one week to perform under ideal conditions, but usually require closer to six to twelve weeks, and even as much as six months. Due to the multiple step reactions involved, these reaction methods produce low yields and are largely unreliable.

One of the primary problems experienced in the art of DNA synthesis is that longer oligonucleotides always have an unavoidable portion of termination products. This is due to the coupling efficiency (how efficiently the DNA synthesizer adds new bases to the growing DNA chain), which only reaches ninety-nine percent per step, even in syntheses that progress well. Deletions may also result from less than one hundred percent capping. With the currently available purification methods, the desired end product can at best obtain ninety-five percent purity. 

Due to the array of potential sequences for even relatively short oligonucleotides, it is also practically impossible to reuse oligonucleotides for various gene constructs, and new oligonucleotides must be synthesized and purified for each new gene construct. Only a fraction of the synthesized material is actually used for the gene synthesis, and the leftover portion is useless. This unsolved incorporation of oligonucleotide synthesis and purification in the gene synthesis process is one of the primary obstacles in obtaining complete automation of the process.

The scientists at Sloning Biotechnology in Denmark sought to overcome these hurdles by developing a method for the efficient synthesis of double-stranded DNA fragments of any sequence and length. The present patent discloses a method that allows any DNA molecule to be constructed from a limited library of basic building blocks. The method allows for parallel synthesis and sequence-independent linkage of any gene fragments. By enacting these two elements, it is possible to achieve a completely automated gene synthesis process.