What is the longest length an oligo can be synthesized?
The real answers lies in the limit of resolution of the purification method and the coupling efficiency of the DNA synthesizer. We can synthesize oligos of 180 bases and obtain sufficient quantities by HPLC purification to do successful gene construction. However, it should be remembered that the longer the oligo, the greater the chance of accumulated sequence errors. Coupling efficiency is the major factor affecting the length of DNA that can be synthesized. Base composition and synthesis scales will also be contributing factors. Table 1 shows that at 99% coupling efficiency, a crude solution of synthesized 95-mers would contain 38% full-length product and 62% (n–x) failure sequences. This is before other chemical effects have been taken into account such as depurination. Depurination mainly affects the base, A. The frequency of depurination is small but will increase significantly with primer length. For these reasons, we specify a maximum length of 80 bases, which we believe is the
