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Abstract As a first step towards the development of radiation hybrid maps, we have produced a radiation hybrid panel in the chicken by fusing female embryonic diploid fibroblasts irradiated at 6 000 rads with HPRT-deficient hamster Wg3hCl2 cells. Due to the low retention frequency of the chicken fragments, a high number of clones was produced from which the best ones were selected. Thus, 452 fusion clones were tested for retention frequencies with a panel of 46 markers. Based on these results, 103 clones with a mean marker retention of 23.8% were selected for large scale culture to produce DNA in sufficient quantities for the genotyping of numerous markers. Retention frequency was tested again with the same 46 markers and the 90 best clones, with a final mean retention frequency of 21.9%, were selected for the final panel. This panel will be a valuable resource for fine mapping of markers and genes in the chicken, and will also help in building BAC contigs.

Abstract Background Owing to the low cost of the high throughput Next Generation Sequencing (NGS) technology, more and more species have been and will be sequenced. However, de novo assemblies of large eukaryotic genomes thus produced are composed of a large number of contigs and scaffolds of medium to small size, having no chromosomal assignment. Radiation hybrid (RH) mapping is a powerful tool for building whole genome maps and has been used for several animal species, to help assign sequence scaffolds to chromosomes and determining their order. Results We report here a duck whole genome RH panel obtained by fusing female duck embryonic fibroblasts irradiated at a dose of 6,000 rads, with HPRT-deficient Wg3hCl2 hamster cells. The ninety best hybrids, having an average retention of 23.6% of the duck genome, were selected for the final panel. To allow the genotyping of large numbers of markers, as required for whole genome mapping, without having to cultivate the hybrid clones on a large scale, three different methods involving Whole Genome Amplification (WGA) and/or scaling down PCR volumes by using the Fluidigm BioMarkTM Integrated Fluidic Circuits (IFC) Dynamic ArrayTM for genotyping were tested. RH maps of APL12 and APL22 were built, allowing the detection of intrachromosomal rearrangements when compared to chicken. Finally, the panel proved useful for checking the assembly of sequence scaffolds and for mapping EST located on one of the smallest microchromosomes. Conclusion The Fluidigm BioMarkTM Integrated Fluidic Circuits (IFC) Dynamic ArrayTM genotyping by quantitative PCR provides a rapid and cost-effective method for building RH linkage groups. Although the vast majority of genotyped markers exhibited a picture coherent with their associated scaffolds, a few of them were discordant, pinpointing potential assembly errors. Comparative mapping with chicken chromosomes GGA21 and GGA11 allowed the detection of the first chromosome rearrangements on microchromosomes between duck and chicken. As in chicken, the smallest duck microchromosomes appear missing in the assembly and more EST data will be needed for mapping them. Altogether, this underlines the added value of RH mapping to improve genome assemblies.

Abstract Background The chicken karyotype is composed of 39 chromosome pairs, of which 9 still remain totally absent from the current genome sequence assembly, despite international efforts towards complete coverage. Some others are only very partially sequenced, amongst which microchromosome 16 (GGA16), particularly under-represented, with only 433 kb assembled for a full estimated size of 9 to 11 Mb. Besides the obvious need of full genome coverage with genetic markers for QTL (Quantitative Trait Loci) mapping and major genes identification studies, there is a major interest in the detailed study of this chromosome because it carries the two genetically independent MHC complexes B and Y. In addition, GGA16 carries the ribosomal RNA (rRNA) genes cluster, also known as the NOR (nucleolus organizer region). The purpose of the present study is to construct and present high resolution integrated maps of GGA16 to refine its organization and improve its coverage with genetic markers. Results We developed 79 STS (Sequence Tagged Site) markers to build a physical RH (radiation hybrid) map and 34 genetic markers to extend the genetic map of GGA16. We screened a BAC (Bacterial Artificial Chromosome) library with markers for the MHC-B, MHC-Y and rRNA complexes. Selected clones were used to perform high resolution FISH (Fluorescent In Situ Hybridization) mapping on giant meiotic lampbrush chromosomes, allowing meiotic mapping in addition to the confirmation of the order of the three clusters along the chromosome. A region with high recombination rates and containing PO41 repeated elements separates the two MHC complexes. Conclusions The three complementary mapping strategies used refine greatly our knowledge of chicken microchromosome 16 organisation. The characterisation of the recombination hotspots separating the two MHC complexes demonstrates the presence of PO41 repetitive sequences both in tandem and inverted orientation. However, this region still needs to be studied in more detail.

Abstract Background Comparative genomics is a powerful means of establishing inter-specific relationships between gene function/location and allows insight into genomic rearrangements, conservation and evolutionary phylogeny. The availability of the complete sequence of the chicken genome has initiated the development of detailed genomic information in other birds including turkey, an agriculturally important species where mapping has hitherto focused on linkage with limited physical information. No molecular study has yet examined conservation of avian microchromosomes, nor differences in copy number variants (CNVs) between birds. Results We present a detailed comparative cytogenetic map between chicken and turkey based on reciprocal chromosome painting and mapping of 338 chicken BACs to turkey metaphases. Two inter-chromosomal changes (both involving centromeres) and three pericentric inversions have been identified between chicken and turkey; and array CGH identified 16 inter-specific CNVs. Conclusion This is the first study to combine the modalities of zoo-FISH and array CGH between different avian species. The first insight into the conservation of microchromosomes, the first comparative cytogenetic map of any bird and the first appraisal of CNVs between birds is provided. Results suggest that avian genomes have remained relatively stable during evolution compared to mammalian equivalents.

Abstract Background The publication of the first draft chicken sequence assembly became available in 2004 and was updated in 2006. However, this does not constitute a definitive and complete sequence of the chicken genome, since the microchromosomes are notably under-represented. In an effort to develop maps for the microchromosomes absent from the chicken genome assembly, we developed radiation hybrid (RH) and genetic maps with markers isolated from sequence currently assigned to "chromosome Unknown" (chrUn). The chrUn is composed of sequence contigs not assigned to named chromosomes. To identify and map sequence belonging to the microchromosomes we used a comparative mapping strategy, and we focused on the small linkage group E26C13. Results In total, 139 markers were analysed with the chickRH6 panel, of which 120 were effectively assigned to the E26C13 linkage group, the remainder mapping elsewhere in the genome. The final RH map is composed of 22 framework markers extending over a 245.6 cR distance. A corresponding genetic map was developed, whose length is 103 cM in the East Lansing reference population. The E26C13 group was assigned to GGA25 (Gallus gallus chromosome 25) by FISH (fluorescence in situ hybridisation) mapping. Conclusion The high-resolution RH framework map obtained here covers the entire chicken chromosome 25 and reveals the existence of a high number of intrachromosomal rearrangements when compared to the human genome. The strategy used here for the characterization of GGA25 could be used to improve knowledge on the other uncharacterized small, yet gene-rich microchromosomes.

Abstract Background By comparing the quail genome with that of chicken, chromosome rearrangements that have occurred in these two galliform species over 35 million years of evolution can be detected. From a more practical point of view, the definition of conserved syntenies helps to predict the position of genes in quail, based on information taken from the chicken sequence, thus enhancing the utility of this species in biological studies through a better knowledge of its genome structure. A microsatellite and an Amplified Fragment Length Polymorphism (AFLP) genetic map were previously published for quail, as well as comparative cytogenetic data with chicken for macrochromosomes. Quail genomics will benefit from the extension and the integration of these maps. Results The integrated linkage map presented here is based on segregation analysis of both anonymous markers and functional gene loci in 1,050 quail from three independent F2 populations. Ninety-two loci are resolved into 14 autosomal linkage groups and a Z chromosome-specific linkage group, aligned with the quail AFLP map. The size of linkage groups ranges from 7.8 cM to 274.8 cM. The total map distance covers 904.3 cM with an average spacing of 9.7 cM between loci. The coverage is not complete, as macrochromosome CJA08, the gonosome CJAW and 23 microchromosomes have no marker assigned yet. Significant sequence identities of quail markers with chicken enabled the alignment of the quail linkage groups on the chicken genome sequence assembly. This, together with interspecific Fluorescence In Situ Hybridization (FISH), revealed very high similarities in marker order between the two species for the eight macrochromosomes and the 14 microchromosomes studied. Conclusion Integrating the two microsatellite and the AFLP quail genetic maps greatly enhances the quality of the resulting information and will thus facilitate the identification of Quantitative Trait Loci (QTL). The alignment with the chicken chromosomes confirms the high conservation of gene order that was expected between the two species for macrochromosomes. By extending the comparative study to the microchromosomes, we suggest that a wealth of information can be mined in chicken, to be used for genome analyses in quail.