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"The conversion from ASCII As, Ts, Gs, and Cs into a stream of bits is done in a fixed-size buffer that assumes a reasonable maximum read length," explained co-author Karl Koscher in response to my requests for more technical information.
The maximum read length of these amplicons is defined by the specifications of GS FLX platform (approximately 240 bases).
This may have important consequences for the development of high-throughput sequencing technologies by suggesting that making high sequencing coverage cheaper may be a better investment of effort than increasing the maximum read length.
Currently, a maximum read length of up to 400 bp can be obtained using this system.
For those CMG genes that were not present in Tier 1 (due to read gaps), we identified those with small (gap < maximum read length) and large gaps (gap > maximum read length).
The reason for this behaviour is that the maximum read length that BSMAP can process is 144 nt.
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In previous studies, PacBio RS data have been reported to contain maximum read lengths of up to 23,000 nucleotides and median lengths of 2,446 nucleotides [ 5].
The Filter FASTQ reads by quality score and length tool allows filtering by minimum and maximum read lengths and by minimum and maximum quality score values over the entire read while allowing a configurable number of deviant bases.
Minimum and maximum read lengths of raw data from SMRT cell 1 were 48 bp and 2,316 bp and for SMRT cell 2 min and max read lengths were 318 bp and 2,364 bp.
All reads where then filtered using pyrocleaner v1.3 (Jérôme et al. 2011) with a quality threshold of 35 and minimum and maximum read lengths set to 100 and 1,000, respectively.
The data produced from the 'third-generation' PacBio RS sequencing platform has a significantly longer read length than that of 'second-generation' technologies such as the Illumina HiSeq2000, and maximum read lengths of 23,000 bp have been reported in the literature, with current average read lengths reaching 2,246 kbp [ 5].
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com