Dataset Information

Construction of papaya male and female BAC libraries and application in physical mapping of the sex chromosomes.

ABSTRACT: Papaya is a major fruit crop in the tropics and has recently evolved sex chromosomes. Towards sequencing the papaya sex chromosomes, two bacterial artificial chromosome (BAC) libraries were constructed from papaya male and female genomic DNA. The female BAC library was constructed using restriction enzyme BstY I and consists of 36,864 clones with an average insert size of 104?kb, providing 10.3x genome equivalents. The male BAC library was constructed using restriction enzyme EcoR I and consists of 55,296 clones with an average insert size of 101?kb, providing 15.0x genome equivalents. The male BAC library was used in constructing the physical map of the male-specific region of the male Y chromosome (MSY) and in filling gaps and extending the physical map of the hermaphrodite-specific region of the Y(h) chromosome (HSY) and the X chromosome physical map. The female BAC library was used to extend the X physical map gap. The MSY, HSY, and X physical maps offer a unique opportunity to study chromosomal rearrangements, Y chromosome degeneration, and dosage compensation of the papaya nascent sex chromosomes.

SUBMITTER: Gschwend AR

PROVIDER: S-EPMC3134383 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Construction of papaya male and female BAC libraries and application in physical mapping of the sex chromosomes.

Gschwend Andrea R AR Yu Qingyi Q Moore Paul P Saski Christopher C Chen Cuixia C Wang Jianping J Na Jong-Kuk JK Ming Ray R

Journal of biomedicine & biotechnology 20110705

Papaya is a major fruit crop in the tropics and has recently evolved sex chromosomes. Towards sequencing the papaya sex chromosomes, two bacterial artificial chromosome (BAC) libraries were constructed from papaya male and female genomic DNA. The female BAC library was constructed using restriction enzyme BstY I and consists of 36,864 clones with an average insert size of 104 kb, providing 10.3x genome equivalents. The male BAC library was constructed using restriction enzyme EcoR I and consists ...[more]

PMID: 21765640

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Project description:BACKGROUND: Papaya is a major fruit crop in tropical and subtropical regions worldwide. It is trioecious with three sex forms: male, female, and hermaphrodite. Sex determination is controlled by a pair of nascent sex chromosomes with two slightly different Y chromosomes, Y for male and Yh for hermaphrodite. The sex chromosome genotypes are XY (male), XYh (hermaphrodite), and XX (female). The papaya hermaphrodite-specific Yh chromosome region (HSY) is pericentromeric and heterochromatic. Physical mapping of HSY and its X counterpart is essential for sequencing these regions and uncovering the early events of sex chromosome evolution and to identify the sex determination genes for crop improvement. RESULTS: A reiterate chromosome walking strategy was applied to construct the two physical maps with three bacterial artificial chromosome (BAC) libraries. The HSY physical map consists of 68 overlapped BACs on the minimum tiling path, and covers all four HSY-specific Knobs. One gap remained in the region of Knob 1, the only knob structure shared between HSY and X, due to the lack of HSY-specific sequences. This gap was filled on the physical map of the HSY corresponding region in the X chromosome. The X physical map consists of 44 BACs on the minimum tiling path with one gap remaining in the middle, due to the nature of highly repetitive sequences. This gap was filled on the HSY physical map. The borders of the non-recombining HSY were defined genetically by fine mapping using 1460?F2 individuals. The genetically defined HSY spanned approximately 8.5?Mb, whereas its X counterpart extended about 5.4?Mb including a 900 Kb region containing the Knob 1 shared by the HSY and X. The 8.5?Mb HSY corresponds to 4.5?Mb of its X counterpart, showing 4?Mb (89%) DNA sequence expansion. CONCLUSION: The 89% increase of DNA sequence in HSY indicates rapid expansion of the Yh chromosome after genetic recombination was suppressed 2-3 million years ago. The genetically defined borders coincide with the common BACs on the minimum tiling paths of HSY and X. The minimum tiling paths of HSY and its X counterpart are being used for sequencing these X and Yh-specific regions.

Project description:BACKGROUND: The papaya Y chromosome has undergone a degenerative expansion from its ancestral autosome, as a consequence of recombination suppression in the sex determining region of the sex chromosomes. The non-recombining feature led to the accumulation of repetitive sequences in the male- or hermaphrodite-specific regions of the Y or the Yh chromosome (MSY or HSY). Therefore, repeat composition and distribution in the sex determining region of papaya sex chromosomes would be informative to understand how these repetitive sequences might be involved in the early stages of sex chromosome evolution. RESULTS: Detailed composition of interspersed, sex-specific, and tandem repeats was analyzed from 8.1 megabases (Mb) HSY and 5.3 Mb corresponding X chromosomal regions. Approximately 77% of the HSY and 64% of the corresponding X region were occupied by repetitive sequences. Ty3-gypsy retrotransposons were the most abundant interspersed repeats in both regions. Comparative analysis of repetitive sequences between the sex determining region of papaya X chromosome and orthologous autosomal sequences of Vasconcellea monoica, a close relative of papaya lacking sex chromosomes, revealed distinctive differences in the accumulation of Ty3-Gypsy, suggesting that the evolution of the papaya sex determining region may accompany Ty3-Gypsy element accumulation. In total, 21 sex-specific repeats were identified from the sex determining region; 20 from the HSY and one from the X. Interestingly, most HSY-specific repeats were detected in two regions where the HSY expansion occurred, suggesting that the HSY expansion may result in the accumulation of sex-specific repeats or that HSY-specific repeats might play an important role in the HSY expansion. The analysis of simple sequence repeats (SSRs) revealed that longer SSRs were less abundant in the papaya sex determining region than the other chromosomal regions. CONCLUSION: Major repetitive elements were Ty3-gypsy retrotransposons in both the HSY and the corresponding X. Accumulation of Ty3-Gypsy retrotransposons in the sex determining region of papaya X chromosome was significantly higher than that in the corresponding region of V. monoica, suggesting that Ty3-Gypsy could be crucial for the expansion and evolution of the sex determining region in papaya. Most sex-specific repeats were located in the two HSY expansion regions.

Project description:BackgroundManduca sexta, Heliothis virescens, and Heliconius erato represent three widely-used insect model species for genomic and fundamental studies in Lepidoptera. Large-insert BAC libraries of these insects are critical resources for many molecular studies, including physical mapping and genome sequencing, but not available to date.ResultsWe report the construction and characterization of six large-insert BAC libraries for the three species and sampling sequence analysis of the genomes. The six BAC libraries were constructed with two restriction enzymes, two libraries for each species, and each has an average clone insert size ranging from 152-175 kb. We estimated that the genome coverage of each library ranged from 6-9 x, with the two combined libraries of each species being equivalent to 13.0-16.3 x haploid genomes. The genome coverage, quality and utility of the libraries were further confirmed by library screening using 6 approximately 8 putative single-copy probes. To provide a first glimpse into these genomes, we sequenced and analyzed the BAC ends of approximately 200 clones randomly selected from the libraries of each species. The data revealed that the genomes are AT-rich, contain relatively small fractions of repeat elements with a majority belonging to the category of low complexity repeats, and are more abundant in retro-elements than DNA transposons. Among the species, the H. erato genome is somewhat more abundant in repeat elements and simple repeats than those of M. sexta and H. virescens. The BLAST analysis of the BAC end sequences suggested that the evolution of the three genomes is widely varied, with the genome of H. virescens being the most conserved as a typical lepidopteran, whereas both genomes of H. erato and M. sexta appear to have evolved significantly, resulting in a higher level of species- or evolutionary lineage-specific sequences.ConclusionThe high-quality and large-insert BAC libraries of the insects, together with the identified BACs containing genes of interest, provide valuable information, resources and tools for comprehensive understanding and studies of the insect genomes and for addressing many fundamental questions in Lepidoptera. The sample of the genomic sequences provides the first insight into the constitution and evolution of the insect genomes.

Project description:BACKGROUND: The genus Aquilegia, consisting of approximately 70 taxa, is a member of the basal eudicot lineage, Ranuculales, which is evolutionarily intermediate between monocots and core eudicots, and represents a relatively unstudied clade in the angiosperm phylogenetic tree that bridges the gap between these two major plant groups. Aquilegia species are closely related and their distribution covers highly diverse habitats. These provide rich resources to better understand the genetic basis of adaptation to different pollinators and habitats that in turn leads to rapid speciation. To gain insights into the genome structure and facilitate gene identification, comparative genomics and whole-genome shotgun sequencing assembly, BAC-based genomics resources are of crucial importance. RESULTS: BAC-based genomic resources, including two BAC libraries, a physical map with anchored markers and BAC end sequences, were established from A. formosa. The physical map was composed of a total of 50,155 BAC clones in 832 contigs and 3939 singletons, covering 21X genome equivalents. These contigs spanned a physical length of 689.8 Mb (~2.3X of the genome) suggesting the complex heterozygosity of the genome. A set of 197 markers was developed from ESTs induced by drought-stress, or involved in anthocyanin biosynthesis or floral development, and was integrated into the physical map. Among these were 87 genetically mapped markers that anchored 54 contigs, spanning 76.4 Mb (25.5%) across the genome. Analysis of a selection of 12,086 BAC end sequences (BESs) from the minimal tiling path (MTP) allowed a preview of the Aquilegia genome organization, including identification of transposable elements, simple sequence repeats and gene content. Common repetitive elements previously reported in both monocots and core eudicots were identified in Aquilegia suggesting the value of this genome in connecting the two major plant clades. Comparison with sequenced plant genomes indicated a higher similarity to grapevine (Vitis vinifera) than to rice and Arabidopsis in the transcriptomes. CONCLUSIONS: The A. formosa BAC-based genomic resources provide valuable tools to study Aquilegia genome. Further integration of other existing genomics resources, such as ESTs, into the physical map should enable better understanding of the molecular mechanisms underlying adaptive radiation and elaboration of floral morphology.

Dataset Information

Construction of papaya male and female BAC libraries and application in physical mapping of the sex chromosomes.

Publications

Construction of papaya male and female BAC libraries and application in physical mapping of the sex chromosomes.

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