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Development and Application of Genomic Resources in Non-model Bird Species
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Understanding the genetic basis of biological processes is a fundamental component of modern ecology and evolutionary biology studies. With the recent advent of next generation sequencing (NGS) technologies, it is now possible to perform large genome and transcriptome projects for ecologically important non-model species. In this thesis, I focused on the development and application of genomic resources of two non-model bird species, the black grouse (Tetrao tetrix) and the great snipe (Gallinago media). Using the chicken genome as a reference, I developed a reference guided NGS pipeline to assemble the complete draft genome of black grouse. The draft genome has a good coverage of the main 29 chromosomes of the chicken genome. The genome was used to develop a vast number of genetic markers. Comparing this genome with that of other species, I identified the genomic regions which were important for the lineage specific evolution of black grouse. I also sequenced and characterised the spleen transcriptome of the black grouse. I identified and validated a large number of gene-based microsatellite markers from the transcriptome and identified and confirmed the expression of immune related genes. Using a similar RNA-Seq approach, I also sequenced the blood transcriptomes of 14 great snipe males with different mating success. I identified genes and single nucleotide polymorphisms (SNPs) which might be related to male mating success in this species, both in terms of gene expression levels and genetic variation structure. For the immunologically important major histocompatibility complex (MHC) gene region of black grouse, I constructed a fosmid library and used it to sequence the complete core MHC region of this species. This resource allowed me to perform a comprehensive comparative genomics analysis of the galliform MHC, by which I found that some genes in this region were affected by selective forces. I was also able to develop a single locus genotyping protocol for the duplicated MHC BLB (class IIB) genes and found that the two black grouse BLB loci followed different evolutionary trajectories. This thesis set an example of developing genomic resources in non-model species and applying them in addressing questions relevant to ecology and evolutionary biology.

Place, publisher, year, edition, pages
Uppsala: Uppsala universitet, 2012. , 49 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 999
Keyword [en]
genome, transcriptome, next generation sequencing, non-model species, bioinformatics, reference guided assembly, RNA-Seq, comparative genomics, gene expression, major histocompatibility complex, single nucleotide polymorphism, microsatellite
National Category
Biological Sciences
Research subject
Biology
Identifiers
URN: urn:nbn:se:uu:diva-183645ISBN: 978-91-554-8540-5 (print)OAI: oai:DiVA.org:uu-183645DiVA: diva2:563641
Public defence
2012-12-14, Lindahlssalen, Norbyvägen 18B, Uppsala, 14:00 (English)
Opponent
Supervisors
Available from: 2012-11-26 Created: 2012-10-31 Last updated: 2013-07-22
List of papers
1. The draft genome of the black grouse (Tetrao tetrix): reference guided assembly and evolutionary implications
Open this publication in new window or tab >>The draft genome of the black grouse (Tetrao tetrix): reference guided assembly and evolutionary implications
(English)Manuscript (preprint) (Other academic)
National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-183637 (URN)
Available from: 2012-10-31 Created: 2012-10-31 Last updated: 2013-09-05
2. Transcriptome sequencing of black grouse (Tetrao tetrix) for immune gene discovery and microsatellite development
Open this publication in new window or tab >>Transcriptome sequencing of black grouse (Tetrao tetrix) for immune gene discovery and microsatellite development
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2012 (English)In: Open Biology, ISSN 2046-2441, E-ISSN 2046-2441, Vol. 2, 120054- p.Article in journal (Refereed) Published
Abstract [en]

The black grouse (Tetrao tetrix) is a galliform bird species that is important forboth ecological studies and conservation genetics. Here, we report the sequencing of the spleen transcriptome of black grouse using 454 GS FLX Titanium sequencing. We performed a large-scale gene discovery analysis with a focus on genes that might be related to fitness in this species and also identified a large set of microsatellites. In total, we obtained 182 179 quality-filtered sequencing reads that we assembled into 9035 contigs. Using these contigs and 15 794 length-filtered (greater than 200 bp) singletons, we identified 7762 transcripts that appear to be homologues of chicken genes. A specific BLAST search with an emphasis on immune genes found 308 homologous chicken genes that have immune function, including ten major histocompatibility complex-related genes located on chicken chromosome 16. We also identified 1300 expressed sequence tag microsatellites and were able to design suitable flanking primers for 526 of these. A preliminary test of the polymorphism of the microsatellites found 10 polymorphic microsatellites of the 102 tested. Genomic resources generated in this study should greatly benefit future ecological, evolutionary and conservation genetic studies on this species.

National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-183642 (URN)10.1098/rsob.120054 (DOI)000307112200005 ()
Available from: 2012-10-31 Created: 2012-10-31 Last updated: 2017-12-07Bibliographically approved
3. Transcriptomic divergence in relation to mating success of lekking great snipes (Gallinago media)
Open this publication in new window or tab >>Transcriptomic divergence in relation to mating success of lekking great snipes (Gallinago media)
Show others...
(English)Manuscript (preprint) (Other academic)
National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-183638 (URN)
Available from: 2012-10-31 Created: 2012-10-31 Last updated: 2013-09-05
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5. Evolutionary history of black grouse major histocompatibility complex class IIB genes revealed through single locus sequence-based genotyping
Open this publication in new window or tab >>Evolutionary history of black grouse major histocompatibility complex class IIB genes revealed through single locus sequence-based genotyping
2013 (English)In: BMC Genetics, ISSN 1471-2156, E-ISSN 1471-2156, Vol. 14, 29- p.Article in journal (Refereed) Published
Abstract [en]

Background: Gene duplications are frequently observed in the Major Histocompatibility Complex (MHC) of many species, and as a consequence loci belonging to the same MHC class are often too similar to tell apart. In birds, single locus genotyping of MHC genes has proven difficult due to concerted evolution homogenizing sequences at different loci. But studies on evolutionary history, mode of selection and heterozygosity correlations on the MHC cannot be performed before it is possible to analyse duplicated genes separately. In this study we investigate the architecture and evolution of the MHC class IIB genes in black grouse. We developed a sequence-based genotyping method for separate amplification of the two black grouse MHC class IIB genes BLB1 and BLB2. Based on this approach we are able to study differences in structure and selection between the two genes in black grouse and relate these results to the chicken MHC structure and organization. Results: Sequences were obtained from 12 individuals and separated into alleles using the software PHASE. We compared nucleotide diversity measures and employed selection tests for BLB1 and BLB2 to explore their modes of selection. Both BLB1 and BLB2 are transcribed and display classic characteristics of balancing selection as predicted for expressed MHC class IIB genes. We found evidence for both intra- and interlocus recombination or gene conversion, as well as indication for positive but differential selection at both loci. Moreover, the two loci appear to be linked. Phylogenetic analyses revealed orthology of the black grouse MHC class IIB genes to the respective BLB loci in chicken. Conclusions: The results indicate that the duplication of the BLB gene occurred before the species divergence into black grouse, chicken and pheasant. Further, we conclude that BLB1 and BLB2 in black grouse are subjected to homogenizing concerted evolution due to interlocus genetic exchange after species divergence. The loci are in linkage disequilibrium, which is in line with the theory of tightly coevolving genes within the MHC under the minimal essential MHC hypothesis. Our results support the conclusion that MHC form and function in birds derived from studies on the domesticated chicken are not artefacts of the domestication process.

National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-183639 (URN)10.1186/1471-2156-14-29 (DOI)000318858800001 ()
Available from: 2012-10-31 Created: 2012-10-31 Last updated: 2017-12-07Bibliographically approved

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