A BAC-based physical map of theDrosophila buzzatiigenome

Abstract

Large-insert genomic libraries facilitate cloning of large genomic regions, allow the construction of clone-based physical maps, and provide useful resources for sequencing entire genomes. Drosophila buzzatii is a representative species of the repleta group in the Drosophila subgenus, which is being widely used as a model in studies of genome evolution, ecological adaptation, and speciation. We constructed a Bacterial Artificial Chromosome (BAC) genomic library of D. buzzatii using the shuttle vector pTARBAC2.1. The library comprises 18,353 clones with an average insert size of 152 kb and an ∼18× expected representation of the D. buzzatii euchromatic genome. We screened the entire library with six euchromatic gene probes and estimated the actual genome representation to be ∼23×. In addition, we fingerprinted by restriction digestion and agarose gel electrophoresis a sample of 9555 clones, and assembled them using FingerPrint Contigs (FPC) software and manual editing into 345 contigs (mean of 26 clones per contig) and 670 singletons. Finally, we anchored 181 large contigs (containing 7788 clones) to the D. buzzatii salivary gland polytene chromosomes by in situ hybridization of 427 representative clones. The BAC library and a database with all the information regarding the high coverage BAC-based physical map described in this paper are available to the research community.

Bottlenecks, population differentiation and apparent selection at microsatellite loci in Australian Drosophila buzzatii

“…Fluorescent in situ hybridization was carried out as in González et al (2000), with detection using anti-digoxigenin antibodies labeled with fluorescein-5-isothiocyanate. Chromosomal localization of the hybridization signals ( Figure 2) was determined using the cytological map of D. buzzatii (González et al, 2005). Microsatellites Db003, Db013, Db034 and Db109 were blasted against the D. mojavensis genome sequence, and localized using data on gene localization in D. buzzatii (Ranz et al, 2003).…”

The 19 Genomes of Drosophila: A BAC Library Resource for Genus-Wide and Genome-Scale Comparative Evolutionary Research

“…Utilization of BAC libraries: Although a few Drosophila BAC libraries have already been reported in the literature (Hoskins et al 2000;Locke et al 2000;Gonzalez et al 2005;Osoegawa et al 2007;Murakami et al 2008), this is the first synthesis and characterization of a comprehensive set of BAC library resources for the genus, which fills a critical void for the Drosophila research community. Hybridization of nine different probes to the full set of libraries demonstrates the feasibility of isolating homologous regions across the entire genus.…”

“…BAC libraries are powerful tools for comparative genome research (Kim et al 1996;Hoskins et al 2000;International Human Genome Mapping Consortium 2000a,b;Locke et al 2000;Osoegawa et al 2000Osoegawa et al , 2001Osoegawa et al , 2004Eichler and Dejong 2002;Gregory et al 2002;Gibbs et al 2003;Krzywinske et al 2004;Gonzalez et al 2005;Ammiraju et al 2006; Drosophila 12 Genomes Consortium 2007; Kim et al 2008;Murakami et al 2008), especially in taxa containing highly repetitive genomes (Havlak et al 2004;Ellison and Shaw 2010;Fang et al 2010). Genome sequences are available for 10 of 19 species for which BAC libraries are constructed, some of which were instrumental in facilitating sequence assemblies (Drosophila 12 Genomes Consortium 2007), and they remain a highpriority resource for improving and finishing several of the low coverage draft genome assemblies.…”

Testing Chromosomal Phylogenies and Inversion Breakpoint Reuse in Drosophila

“…Pools were designed to close the gaps between clones already hybridized. BAC clones were chosen according to their localization in the D. buzzatii genome map (González et al 2005) to optimize coverage of chromosomes X (57 clones) and 2 (123 clones). Particular attention was paid to those regions that have been rearranged during the divergence between D. repleta and D. buzzatii.…”

“…These two inversions are apparently fixed in the lineage leading from Primitive I to D. repleta, yet no attempt was made to determine their distribution in the phylogenetic tree (Ranz et al 2003). In a further effort to determine the precise number and extent of structural changes between D. buzzatii and D. repleta, we have turned to clones from the recently produced BAC library and BAC-based physical map of the D. buzzatii genome (González et al 2005). We have mapped to D. repleta chromosomes 180 BAC clones from D. buzzatii chromosomes X and 2 that harbor most of the rearrangements fixed between these two species ( Figure 1A).…”