Theories and applications for sequencing randomly selected clones

Theory is developed for the process of sequencing randomly selected large-insert clones. Genome size, library depth, clone size, and clone distribution are considered relevant properties and perfect overlap detection for contig assembly is assumed. Genome-specific and nonrandom effects are neglected. Order of magnitude analysis indicates library depth is of secondary importance compared to the other variables, especially as clone size diminishes. In such cases, the well-known Poisson coverage law is a good approximation. Parameters derived from these models are used to examine performance for the specific case of sequencing random human BAC clones. We compare coverage and redundancy rates for libraries possessing uniform and nonuniform clone distributions. Results are measured against data from map-based human-chromosome-2 sequencing. We conclude that the map-based approach outperforms random clone sequencing, except early in a project. However, simultaneous use of both strategies can be beneficial if a performance-based estimate for halting random clone sequencing is made. Results further show that the random approach yields maximum effectiveness using nonbiased rather than biased libraries.