Remarkably, the use of reads of only 100 bp had a relatively small effect on the mean error of reassembly (Figure 5B), compared to the error when reads of
150 bp were used (Figure 5A). For sets of reassemblies with reads of 150 bp there was little change in the error for fragments between 100 and 600 kb in length. Each data point is an average over 27 repeated simulations. However, for reads of 100 bp, although error remained constant for 10 mutants and 10fold coverage over the entire 600 kb, there was a trebling of the error for lengths longer than 300 kb when 10 mutants at 8-fold coverage were analysed. The sudden increase in the proportion of errors between 200 and 300 kb fragments is probably due to the presence of reconstruction ambiguities in this part of the fragment. That this problem is not seen for reads of length 150 bp may indicate that the ambiguities are resolved for longer reads. Thus, it also appears that increasing the level of fragment coverage can improve the reconstruction of individual mutants at stage I, whereas analysis of additional mutants may reduce the proportion of error. The number of mutants necessary for error-free reconstruction of these three human genomic regions using 150 bp reads is shown in Figure 6. These data suggest that high levels of coverage (c — 10) and use of either 12 or 15 mutants allow reconstruction with proportions of errors less than 1/10,000. High-density PicoTiterPlate sequencing arrays (Margulies et al., 2005, 2007) and polony arrays (Shendure et al, 2005) can readily achieve these levels of sequencing coverage within a single experiment.
effect of mutant number
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