regions outside this ‘safe’ range will suffer substantial G+C biases. The G+C% bias appears asymmetric, with the bias against high G+C% regions (i.e. D. maricopensis) more pronounced than low G+C% regions (i.e. S. tokodaii). In fact this bias against extremely high G+C% templates may explain why sequencing D. maricopensis libraries, as well as Streptomyces sp. and Burkholderia sp. libraries, either failed during library preparation or produced insufficient sequence data. PGM sequencing of organisms with low or high G+C% will experience substantial read-depth biases, however, more concerning is this failure to produce libraries from high G+C% templates. This bias will likely result in inaccurate representation of organisms in metagenomic and metatranscriptomic data generated on the PGM.
