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Fosmid Libraries for Genomic Structural Variation Detection

2007; Wiley; Volume: 54; Issue: 1 Linguagem: Inglês

10.1002/0471142905.hg0520s54

1934-8266

William F. Donahue, Heather Ebling,

Chromosomal and Genetic Variations

Fosmid libraries have demonstrated their utility for a number of applications. These include filling gaps between BACs and small insert libraries in sequence assemblies, performing hybridization/screening studies to isolate functional elements within the genome (Vergin et al., 1998), and detecting insertions, deletions, and rearrangements in structural variation studies (Tuzun et al., 2005). This unit covers the basic methodologies for the construction of fosmid libraries with tight insert sizes suitable for these applications. Basic Protocol 1 covers the shearing, size selection, and recovery of DNA from a pulsed-field gel. Basic Protocol 2 covers the cloning of insert DNA into the fosmid vector, packaging of DNA into infective phage particles, and the infection/transformation of bacteria. A commentary section is provided, which outlines many of the critical parameters involved in fosmid library construction, along with some additional background information and a section discussing anticipated results.