A SNP based linkage map of the turkey genome reveals multiple intrachromosomal rearrangements between the Turkey and Chicken genomes

The turkey (Meleagris gallopavo, MGA) is an important agricultural species that is largely used as a meat type bird. In 2008, turkey represented 6.65% of the world poultry meat production [1]. The world-wide turkey population has rapidly grown due to increased commercial farming. Global turkey stocks nearly tripled from 178 million in 1970 to over 482 million in 2008. Over the same time period, the production volume increased more than fivefold from 1.2 to 6.1 million tons [1].

The turkey genome consists of 39 pairs of autosomes and 1 pair of sex chromosomes [2]. The predicted size of the turkey genome is 1.1 billion bases on the turkey genome build UMD 2.01 which is based on sequences from a combination of two next generation sequencing platforms, Roche 454 and Illumina GAII with 5× and 25× coverage respectively. Limited information is available on the turkey linkage and physical map although a small number of low resolution linkage maps using microsatellite markers [2, 3] have been published. Linkage maps of chicken on the other hand are more abundant and have generally used larger numbers of markers [4–10]. Comparative cytogenetic and linkage maps between turkey and chicken showed conserved synteny and close ancestral relation among these species [2, 3] and support the hypothetical ancestral Galliform karyotype [11]. Chromosome banding and zoo-FISH with chromosome paints for the turkey and chicken chromosomes have suggested that chicken and turkey karyotypes are distinguished by at least two interchromosomal rearrangements [2, 12, 13]. Chicken chromosome 2 and 4 are represented by turkey chromosomes 3 and 6 and by turkey chromosomes 4 and 9 respectively [2, 3, 13].

Chromosome studies have revealed that the karyotype is more conserved among avian lineages than it is among other groups, such as mammals, with most avian species showing a diploid chromosome number between 76 and 80 http://www.genomesize.com. This suggests that chromosomal evolution or large-scale rearrangements affecting chromosome number occur at a low rate in birds and as a result many chromosomes have remained more or less intact during avian evolution [14]. Chicken chromosome specific probes have been used for in situ hybridization onto metaphase spreads of other birds and revealed an overall picture of a high degree of chromosomal homology between chicken and representatives from many avian orders [15]. Hybridization results also indicated that interchromosomal rearrangements have been rare during avian evolution [16, 17].

A linkage map is essential for the mapping of quantitative trait loci (QTL) and very useful for the assembly of genome sequence and subsequently mapping of genes along the chromosomes. A high-resolution linkage map facilitates fine mapping of quantitative trait loci (QTLs) and can be produced because of the abundance of SNPs within the genome [18]. SNP based genotyping is preferred because it is highly accurate, quick and automated, using limited human intervention. Increasing the marker density of the linkage map further enables the analyses of genomic sequences associated with high recombination rates [9].

The present study was designed to develop a SNP based linkage map in turkey and to detect genomic rearrangements between turkey and chicken.

A whole genome SNP-based linkage map for the turkey is presented with 531 markers dispersed over 28 linkage groups and a total map length of 2324 cM. The total map length in the present study was slightly higher than that described by Reed et al. [3]. This difference in length is probably caused by coverage of three additional turkey chromosomes (MGA20, 24 and MGA25) along with the utilization of 69% higher number of markers in the present study, likely to be covering a larger proportion of the turkey genome. Matching the turkey linkage groups with the chicken physical map identified a map for each of the syntenic groups/chromosomes described by Griffin et al. [2]. The comparison of the syntenic chromosomes between turkey and chicken showed that the genetic lengths of turkey chromosomes were very similar to the estimated genetic length of the chicken chromosomes (Table 1). When comparing the turkey genetic linkage map and chicken physical map with respect to the order of markers across the chromosomes, some of the chromosomes (MGA14, 21, 25, 26 and MGAZ) showed complete conservation in the order of markers whereas others showed limited variation. The conservation in the order of markers for the chicken chromosomes GGA12, 19, 24 and GGAZ with the syntenic turkey linkage groups was also observed by Reed et al. [3]. This high rate of concordance in the order of markers between the genomes of these two avian species is indicative of a highly conserved nature of avian genomes.

Observed interchromosomal rearrangements (Figure 1) in the present study between turkey and chicken are in agreement with the results of Griffin et al. [2]. A number of complex intrachromosomal rearrangements (inversions) were also observed between turkey and chicken. The observed large inverted regions, of nearly a complete chromosome arm on MGA10 and MGA20 in comparison to their syntenic chicken chromosome GGA8 and GGA18 (Figure 2) were also observed in a sequence based comparative study by Dalloul et al. [29]. Cytogenetic studies using chromosome painting also reported an inversion on MGA10 in comparison to the syntenic chicken chromosome GGA8 [2, 13]. Our comparative linkage map of turkey and chicken does not show pericentic inversions on MGA2 and MGA3p as were reported by Griffin et al. [2] but we have observed complex rearrangements resulting in a reversed order of markers on these chromosomes (Additional file 3). Several other chromosomes, notably MGA1, 2, 5, 8, 11, 12, 13, 15, 16, 17, 19, 22, and MGA28 as well as the chromosomes that showed interchromosomal rearrangements (MGA3 and MGA6; MGA4 and MGA9) between turkey and chicken, also showed additional complex rearrangements probably involving multiple inversions or other complex rearrangements (Figure 1). A higher number of rearrangements per Mb were observed on the microchromosomes than on the macrochromosomes. The occurrence of this high number of rearrangements at the microchromosomes could be explained by the positive association of rearrangements with recombination rate [30].

Our observed low number of interchromosomal rearrangements between the chicken and turkey genomes, confirms previous results of a high degree of interchromosomal synteny in birds as seen within a number of different comparative studies of chicken with quail, duck and zebra finch [30–32]. It has been suggested that the low number of interchromosomal rearrangements during avian genome evolution is a consequence of the small amount of interspersed repeats, segmental duplications, and pseudogenes in avian genomes, which provide little opportunity for non allelic homozygous recombination [33, 34]. A relatively high number of intrachromosomal rearrangements was observed in our comparative analysis of the turkey and chicken, which agrees with the findings of the sequence based comparative studies of chicken with turkey and zebra finch [26, 29, 30]. The relatively high number of intrachromosomal rearrangements clearly suggests that the organization of avian genomes is more prone to intrachromosomal rearrangements than previously appreciated based on chromosome banding and chromosome painting data [2].

The comparison of male vs. female maps showed differences in genetic lengths of maps. In turkey, the total male-specific map appeared to be 195 cM longer than the female specific map. However, female-specific maps for some chromosomes (MGA10, 11, 15, 16, 23, 25, 26 and MGA28) were also found to be longer than the male maps (Table 1). The longer map length in homogametic males can be explained by the Haldane-Huxley rule [35, 36], which predicts that the frequency of recombination during meiosis is lower in the heterogametic sex. The smaller map lengths in turkey for some male-specific maps were found to be an exception to the Haldane-Huxley rule. However, the longer map lengths for some chromosome maps in the heterogametic sex were also found in chicken [9].

In the present study three maps i.e. the turkey genetic linkage map, the turkey physical map and the chicken physical map were compared. The discordance of turkey physical map with the turkey genetic linkage and the chicken physical map in the allocation of marker at different chromosomes could possibly be explained by the occurrence of assembly errors in the turkey genome sequence. The turkey physical map was created completely by whole genome shotgun sequencing using Roche 454 and Illumina GA2 sequence data. Inconsistencies between the turkey linkage and chicken physical maps relative to the turkey physical map are most likely a reflection of the challenge of correctly assembling a genome based on next-gen sequencing data alone. Markers that were in agreement between turkey linkage and chicken physical maps but that could not be positioned on the turkey physical map most likely reflect an uncovered genomic regions since the turkey genome sequence is known to cover around 95% of the complete genome (Turkey genome build UMD 2.01).

In general, higher recombination rates and higher densities of GC-rich elements were found on microchromosomes compared to macrochromosomes (Figure 3A & 3B). During meiosis, at least one chiasma per bivalent chromosome is required [37], but the likelihood of chiasmata forming varies along the chromosome [38]. In turkey, recombination rate and GC rich sequences (CTCF, CCTCCCT, CTCTCCC and CCCCCCC) were found to co-vary among different chromosomes. A similar trend was also seen in human, mouse and other birds like chicken and zebra finch [9, 10, 20, 26, 37, 39, 40].

In the present study recombination rates were found to be correlated with CpG/Mb. In general CpG/Mb tended to increase in areas of higher recombination i.e. microchromosomes (Figure 3B). This demonstrates that in the turkey microchromosomes, high recombination rate, high amount of GC-rich sequences (CTCF, CCTCCCT, CTCTCCC and CCCCCCC) and high amount of CpG contents are all correlated (Figure 3A-F). Other studies reported that GC-rich regions in a genome had higher gene densities [41, 42] and that microchromosomes had higher gene densities than the macrochromosomes [38]. The nature of the microchromosomes in birds, with their high recombination rates, high amount of GC-rich sequences, GC content and gene densities appears to be an extreme instance of a general trend.

The results for MGAZ in the analysis of recombination rate and sequence motif densities across the chromosomes, were unexpected and MGAZ appeared as outlier as seen in figure 3A-F. This outlier spot could represent a true characteristic of MGAZ but more likely results from the low marker density on this particular chromosome in our analysis. (Additional Files 2 & 3).

Our SNP-based genetic linkage map of turkey with 531 markers reveals a genome length of 2,324 cM. This linkage map also allowed a comparison of the genome structures of turkey and chicken, demonstrating a very high degree of conservation in chromosome structure. A relatively low number of inter-, and intrachromosomal rearrangements was observed despite these two species being separated by 40 million years of evolution.