Bacterial artificial chromosome libraries for mouse sequencing and functional analysis.

作者：Kazutoyo Osoegawa, Morihiro Tateno, Peng Yeong Woon, Eirik Frengen, Aaron Gerald Mammoser, Joseph J. Catanese, Yoshihide Hayashizaki, Pieter J. de Jong · 发表于：PubMed · 年份：2000 · 被引用次数：302 · 研究领域：Genomics and Phylogenetic Studies、Chromosomal and Genetic Variations、Glycosylation and Glycoproteins Research

Bacterial artificial chromosome (BAC) and P1-derived artificial chromosome (PAC) libraries providing a combined 33-fold representation of the murine genome have been constructed using two different restriction enzymes for genomic digestion. A large-insert PAC library was prepared from the 129S6/SvEvTac strain in a bacterial/mammalian shuttle vector to facilitate functional gene studies. For genome mapping and sequencing, we prepared BAC libraries from the 129S6/SvEvTac and the C57BL/6J strains. The average insert sizes for the three libraries range between 130 kb and 200 kb. Based on the numbers of clones and the observed average insert sizes, we estimate each library to have slightly in excess of 10-fold genome representation. The average number of clones found after hybridization screening with 28 probes was in the range of 9-14 clones per marker. To explore the fidelity of the genomic representation in the three libraries, we analyzed three contigs, each established after screening with a single unique marker. New markers were established from the end sequences and screened against all the contig members to determine if any of the BACs and PACs are chimeric or rearranged. Only one chimeric clone and six potential deletions have been observed after extensive analysis of 113 PAC and BAC clones. Seventy-one of the 113 clones were conclusively nonchimeric because both end markers or sequences were mapped to the other confirmed contig members. We could not exclude chimerism for...