Genome assembly. chromosomal organization and comparative genomics of multiple bird species: beyond "catalogues of genes"

Lead Research Organisation: University of Kent
Department Name: Sch of Biosciences

Abstract

Imagine trying to navigate through a town with only the index from your A-Z to guide you and not the street map. In essence, this is what happens when the genome of a new species is sequenced. Old fashioned technology used to "sketch the outline of the streets" first before filling in the gaps with the gene sequence. The new technology (so called "Next Generation Sequencing") is much cheaper and quicker but is quite poor at sorting out where the genes are in relation to one another. Essentially, Next Generation Sequencing assemblies are little more than catalogues of genes with little structure of the overall genome apparent. Fortunately this problem can be rectified using a technique called "FISH" that can take the data from next generation sequencing projects and visualise directly the genes as they appear in their rightful place in the genome. In this project we intend to do this with 25-30 recently completed genome sequences from birds. Birds are among the most diverse animals with around 10,000 living species. Many are models for human disease and development and are critical to agriculture (both meat and eggs). Others are threatened or endangered and, with impending global warming, molecular tools for the study of ecology and conservation of birds are essential. It is also possible to compare the overall structure of one genome with another, somewhat like comparing the maps of towns with similar layouts. We have developed online tools that can directly visualize the similarities and differences between the genomes of several birds at a time. Of course the differences between these bird genomes came about through changes that happened during evolution. One of the main aims of this project is to find out how and why this occurred. We have a number of ideas such as we think there may be different "gene signatures" at the places in the genome where evolutionary rearrangements are more or less likely to occur. We think that the whole process may be related to the mechanisms by which our genes recombine in our germ cells. We also think there may be a role for small pieces of DNA that "float around" these bird genomes. Finally, we have received overwhelming support from numerous laboratories all over the world, all of whom are interested in bird genomics and who would wish to use our online resource to ask biological questions of their own. For this reason we feel that this project not only will help us understand evolution in birds but also establish the UK as a central international hub for work of this kind.

Technical Summary

Every genome sequence needs a good map however modern next generation sequencing (NGS) technologies struggle fully to assemble whole genomes de-novo. This is disappointing as mapping information on a chromosomal basis brings the opportunity to address many fundamental biological questions of genome evolution. Modern genomic research generally focuses on the role of individual genes but the role of chromosomes and homologous synteny blocks (HSBs) is often disregarded. This is despite the ubiquity of HSBs and the phenotypic consequences/evolutionary implications of chromosome rearrangement. In mammals, there are sufficient assembled reference genomes to make assembly by comparative analysis. For birds however only three such reference genomes exist and chromosomal assembly needs to be achieved by other means. Classical approaches involving karyotyping and FISH coupled with comparative genomics browser technology provide a straightforward solution for the many avian genomes that are assembled to the point of 100+ large scaffolds. The aim of the project will thus be to generate a freely available comparative genomic resource that can be used to compare the genome structures of 25-30 bird species. HSBs and Evolutionary Breakpoint Regions (EBRs) will be displayed using the "Evolution Highway" browser. We will anchor each of the scaffolds physically to chromosomes by FISH. We will address at least four fundamental questions pertaining to genome evolution: We will test the hypotheses that HSB lengths are non-randomly distributed in birds, that there are specific gene ontology signatures of HSBs and EBRs, that EBRs coincide with recombination hotspots in birds (but not in mammals) and that transposable element density in increased in EBRs. The extensive letters of support from potential users indicate that this will be a widely used resource addressing a myriad of further biological questions in this most diverse and biologically important group of vertebrates.

Planned Impact

Exploitation and Application.
Technology Transfer activity at both Universities is administered by Enterprise offices. Collaborative agreement already exist with Digital Scientific and CytoCell. These separately capture exploitation and collaborative output issues going forward. Cytocell, Aviagen and Digital Scientific UK see benefit to this project either through the identification of commercially important traits or through the sales of products and have thus offered generous in kind support. A specific strategic advisory board has been established to identify potential impact issues from this and other projects.

Communications and engagement with the identified beneficiaries.
The main beneficiaries of this grant will be the avian genomics community in industry, third sector and academia. In 2002 the Kent lab was set up as a resource centre for avian molecular cytogenetics (FARMACRHOM). Through these activities we developed a mature and functioning collaborative network with most of the major players in avian genomics and biology many of whom have attached letters of support. Evolution Highway the "industry standard" for browser comparative genomics in the mammalian chromosome world with over ~50 regular users 16,000 hits from academia, industry and third sector in the last year. Inherently used for collaborative research it is, in itself, a tool to facilitate communication and engagement. Another beneficiary is the poultry breeding industry through which the Kent lab is, and has been, supported and funded through BBSRC grants and PhD CASE studentships. In the past we have provided genome maps for the species of interest (turkey and duck). A local zoo (Wingham Wildlife Park) was engaged as another and benefitted by subsequent broadcast media coverage and by provision of a display on the collaborative work performed (this is currently on display to the general public). We will participate in company-managed events and community activities with specialist audiences and the general public to disseminate findings and will develop as a CPD module for delivery to outside engagers. We will contribute to the Bioscience KTN newsletter, to "BBSRC business," newsletters published by individual companies and by the Departments/Universities and to media coverage as well as online activities. Bioscience KTN provide further help and advice on collaboration and their website has >3,000 registered members working in industry and academia The PI's have a healthy relationship with the University press offices and are regular contributor to public engagement activities including "Café Scientifique," Science Fairs and public lectures. Thus far, this work has led to several press releases in addition to appearances in the national and local broadcast media (BBC's "Big Questions," "The World Tonight", Radio 2 Simon Mayo show" BBC radio Kent etc.). Both Universities have active outreach programmes to which the PIs will contribute.

Capacity and Involvement
The two PIs will be the primary participants in impact activities aided by by PhD students and post-doctoral researchers. Professor Griffin has extensive training and experience in communication activities and commercial exploitation of research. The communications and development office in both Universities provide technical expertise and help in writing publications, web pages and user-friendly interfaces and it is expected that post-docs and PhD students will be provided with significant training in both dissemination and exploitation activities through Departmental, University and Bioscience KTN related activities. In conclusion therefore, this project has a range of expertise and ambitious, but achievable plans for impact at local, regional national and international levels.

See complete impact plan (attached) for further details

Publications

10 25 50

publication icon
Lithgow PE (2014) Novel tools for characterising inter and intra chromosomal rearrangements in avian microchromosomes. in Chromosome research : an international journal on the molecular, supramolecular and evolutionary aspects of chromosome biology

 
Title Additional file 1 of Analysis of multiple chromosomal rearrangements in the genome of Willisornis vidua using BAC-FISH and chromosome painting on a supposed conserved karyotype 
Description Addtional file 1. Phylogenetic relationship among chicken, zebra finch, the Eurasian Stone Curlew and the Wedge-Billed Woodcreeper. The phylogeny is based in Prum et al. [48]. 
Type Of Art Film/Video/Animation 
Year Produced 2021 
URL https://springernature.figshare.com/articles/figure/Additional_file_1_of_Analysis_of_multiple_chromo...
 
Title Additional file 1 of Analysis of multiple chromosomal rearrangements in the genome of Willisornis vidua using BAC-FISH and chromosome painting on a supposed conserved karyotype 
Description Addtional file 1. Phylogenetic relationship among chicken, zebra finch, the Eurasian Stone Curlew and the Wedge-Billed Woodcreeper. The phylogeny is based in Prum et al. [48]. 
Type Of Art Film/Video/Animation 
Year Produced 2021 
URL https://springernature.figshare.com/articles/figure/Additional_file_1_of_Analysis_of_multiple_chromo...
 
Title Additional file 1: of Chromosome map of the Siamese cobra: did partial synteny of sex chromosomes in the amniote represent "a hypothetical ancestral super-sex chromosome" or random distribution? 
Description Figure S1. Chromosomal locations of microsatellite repeat motifs in the Siamese cobra (Naja kaouthia). Hybridization patterns of FITC-labeled (CA)15 (a), (AT)15 (b), (GC)15 (c), (CAT)10 (d), and (AAT)10 (e) on DAPI-stained chromosomes. Scale bar represents 10 µm. (JPG 175 kb) 
Type Of Art Film/Video/Animation 
Year Produced 2018 
URL https://springernature.figshare.com/articles/Additional_file_1_of_Chromosome_map_of_the_Siamese_cobr...
 
Title Additional file 1: of Chromosome map of the Siamese cobra: did partial synteny of sex chromosomes in the amniote represent "a hypothetical ancestral super-sex chromosome" or random distribution? 
Description Figure S1. Chromosomal locations of microsatellite repeat motifs in the Siamese cobra (Naja kaouthia). Hybridization patterns of FITC-labeled (CA)15 (a), (AT)15 (b), (GC)15 (c), (CAT)10 (d), and (AAT)10 (e) on DAPI-stained chromosomes. Scale bar represents 10 µm. (JPG 175 kb) 
Type Of Art Film/Video/Animation 
Year Produced 2018 
URL https://springernature.figshare.com/articles/Additional_file_1_of_Chromosome_map_of_the_Siamese_cobr...
 
Title Additional file 2: of Chromosome map of the Siamese cobra: did partial synteny of sex chromosomes in the amniote represent â a hypothetical ancestral super-sex chromosomeâ or random distribution? 
Description Figure S2. Cytogenetic map of the Siamese cobra (Naja kaouthia), which shows chromosome homologies with chicken and zebra finch. This map was constructed with 25 chicken and zebra finch BACs mapped on the Siamese cobra macrochromosomes. Locations of BACs are shown to the right of the Siamese cobra chromosomes. The chromosome numbers show the chromosomes of the chicken (Gallus gallus, GGA) and zebra finch (Taeniopygia guttata, TGU), which show homologies with the Siamese cobra chromosomes. (JPG 118 kb) 
Type Of Art Film/Video/Animation 
Year Produced 2018 
URL https://springernature.figshare.com/articles/Additional_file_2_of_Chromosome_map_of_the_Siamese_cobr...
 
Title Additional file 2: of Chromosome map of the Siamese cobra: did partial synteny of sex chromosomes in the amniote represent â a hypothetical ancestral super-sex chromosomeâ or random distribution? 
Description Figure S2. Cytogenetic map of the Siamese cobra (Naja kaouthia), which shows chromosome homologies with chicken and zebra finch. This map was constructed with 25 chicken and zebra finch BACs mapped on the Siamese cobra macrochromosomes. Locations of BACs are shown to the right of the Siamese cobra chromosomes. The chromosome numbers show the chromosomes of the chicken (Gallus gallus, GGA) and zebra finch (Taeniopygia guttata, TGU), which show homologies with the Siamese cobra chromosomes. (JPG 118 kb) 
Type Of Art Film/Video/Animation 
Year Produced 2018 
URL https://springernature.figshare.com/articles/Additional_file_2_of_Chromosome_map_of_the_Siamese_cobr...
 
Description Throughout the course of this project, we have make substantial progress with numerous publications. We have mapped the genomes of several species as well as provided insight into the genomes of dinosaurs
Exploitation Route The project is ongoing and we have numerous collaborators
Sectors Agriculture

Food and Drink

Education

URL http://www.bbsrc.ac.uk/news/fundamental-bioscience/2017/170105-pr-complete-animal-genomes-become-easier-to-map/
 
Description I continue to get several invitations to give public and academic lectures based around the work. We have worked with a company Cytocell in order to develop screening devices for infertility and comparative genomics in the agricultural industry. We now market these devices and are spinning off a company to progress this further
First Year Of Impact 2017
Sector Agriculture, Food and Drink,Education
Impact Types Cultural

 
Description Rapid reconstruction of reference chromosome-level mammalian genome assemblies and insight into the mechanisms of gross genomic rearrangement
Amount £292,975 (GBP)
Funding ID BB/P020054/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 09/2017 
End 09/2021
 
Title Additional file 1: of Chromosome-level assembly reveals extensive rearrangement in saker falcon and budgerigar, but not ostrich, genomes 
Description Intrachromosomal rearrangements: BAC IDs and chromosomal orientation of clones (with start and stop coordinates from the chicken genome). The order of clones from the top to the bottom represents the order in which that appears on the chromosomes of the species of interest. Text in red indicates the p- (short) arm of the chromosome (where it is discernable). Data is listed in supplementary tables as follows: Table S1. Ostrich genome; Table S2. Budgerigar genome; Table S3. Saker falcon genome. (ZIP 58 kb) 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
URL https://springernature.figshare.com/articles/Additional_file_1_of_Chromosome-level_assembly_reveals_...
 
Title Additional file 1: of Chromosome-level assembly reveals extensive rearrangement in saker falcon and budgerigar, but not ostrich, genomes 
Description Intrachromosomal rearrangements: BAC IDs and chromosomal orientation of clones (with start and stop coordinates from the chicken genome). The order of clones from the top to the bottom represents the order in which that appears on the chromosomes of the species of interest. Text in red indicates the p- (short) arm of the chromosome (where it is discernable). Data is listed in supplementary tables as follows: Table S1. Ostrich genome; Table S2. Budgerigar genome; Table S3. Saker falcon genome. (ZIP 58 kb) 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
URL https://springernature.figshare.com/articles/Additional_file_1_of_Chromosome-level_assembly_reveals_...
 
Title Additional file 2: of Chromosome-level assembly reveals extensive rearrangement in saker falcon and budgerigar, but not ostrich, genomes 
Description Chromosomal coordinates and orientation of mapped scaffolds and PCFs are listed by chromosome for each species. Data is listed in supplementary tables as follows: Table S4. Ostrich genome; Table S5. Budgerigar genome; Table S6. Saker falcon genome. (ZIP 87 kb) 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
URL https://springernature.figshare.com/articles/Additional_file_2_of_Chromosome-level_assembly_reveals_...
 
Title Additional file 2: of Chromosome-level assembly reveals extensive rearrangement in saker falcon and budgerigar, but not ostrich, genomes 
Description Chromosomal coordinates and orientation of mapped scaffolds and PCFs are listed by chromosome for each species. Data is listed in supplementary tables as follows: Table S4. Ostrich genome; Table S5. Budgerigar genome; Table S6. Saker falcon genome. (ZIP 87 kb) 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
URL https://springernature.figshare.com/articles/Additional_file_2_of_Chromosome-level_assembly_reveals_...
 
Title Additional file 3: of Chromosome-level assembly reveals extensive rearrangement in saker falcon and budgerigar, but not ostrich, genomes 
Description EBRs detected and genome position in relation to the chicken genome. Data is listed in supplementary tables as follows: Table S7. Ostrich genome; Table S8. Budgerigar genome; Table S9. Saker falcon genome. (ZIP 61 kb) 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
URL https://springernature.figshare.com/articles/Additional_file_3_of_Chromosome-level_assembly_reveals_...
 
Title Additional file 3: of Chromosome-level assembly reveals extensive rearrangement in saker falcon and budgerigar, but not ostrich, genomes 
Description EBRs detected and genome position in relation to the chicken genome. Data is listed in supplementary tables as follows: Table S7. Ostrich genome; Table S8. Budgerigar genome; Table S9. Saker falcon genome. (ZIP 61 kb) 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
URL https://springernature.figshare.com/articles/Additional_file_3_of_Chromosome-level_assembly_reveals_...
 
Title Cytotaxonomy of Gallinula melanops (Gruiformes, Rallidae): Karyotype evolution and phylogenetic inference 
Description Abstract Although Rallidae is the most diverse family within Gruiformes, there is little information concerning the karyotype of the species in this group. In fact, Gallinula melanops, a species of Rallidae found in Brazil, is among the few species studied cytogenetically, but only with conventional staining and repetitive DNA mapping, showing 2n=80. Thus, in order to understand the karyotypic evolution and phylogeny of this group, the present study aimed to analyze the karyotype of G. melanops by classical and molecular cytogenetics, comparing the results with other species of Gruiformes. The results show that G. melanops has the same chromosome rearrangements as described in Gallinula chloropus (Clade Fulica), including fission of ancestral chromosomes 4 and 5 of Gallus gallus (GGA), beyond the fusion between two of segments resultants of the GGA4/GGA5, also fusions between the chromosomes GGA6/GGA7. Thus, despite the fact that some authors have suggested the inclusion of G. melanops in genus Porphyriops, our molecular cytogenetic results confirm its place in the Gallinula genus. 
Type Of Material Database/Collection of data 
Year Produced 2022 
Provided To Others? Yes  
URL https://scielo.figshare.com/articles/dataset/Cytotaxonomy_of_Gallinula_melanops_Gruiformes_Rallidae_...
 
Title Cytotaxonomy of Gallinula melanops (Gruiformes, Rallidae): Karyotype evolution and phylogenetic inference 
Description Abstract Although Rallidae is the most diverse family within Gruiformes, there is little information concerning the karyotype of the species in this group. In fact, Gallinula melanops, a species of Rallidae found in Brazil, is among the few species studied cytogenetically, but only with conventional staining and repetitive DNA mapping, showing 2n=80. Thus, in order to understand the karyotypic evolution and phylogeny of this group, the present study aimed to analyze the karyotype of G. melanops by classical and molecular cytogenetics, comparing the results with other species of Gruiformes. The results show that G. melanops has the same chromosome rearrangements as described in Gallinula chloropus (Clade Fulica), including fission of ancestral chromosomes 4 and 5 of Gallus gallus (GGA), beyond the fusion between two of segments resultants of the GGA4/GGA5, also fusions between the chromosomes GGA6/GGA7. Thus, despite the fact that some authors have suggested the inclusion of G. melanops in genus Porphyriops, our molecular cytogenetic results confirm its place in the Gallinula genus. 
Type Of Material Database/Collection of data 
Year Produced 2022 
Provided To Others? Yes  
URL https://scielo.figshare.com/articles/dataset/Cytotaxonomy_of_Gallinula_melanops_Gruiformes_Rallidae_...
 
Title Data from: Chromosome-level assembly reveals extensive rearrangement in saker falcon and budgerigar, but not ostrich, genomes 
Description The number of de novo genome sequence assemblies is increasing exponentially; however, relatively few contain one scaffold/contig per chromosome. Such assemblies are essential for studies of genotype-to-phenotype association, gross genomic evolution, and speciation. Inter-species differences can arise from chromosomal changes fixed during evolution, and we previously hypothesized that a higher fraction of elements under negative selection contributed to avian-specific phenotypes and avian genome organization stability. The objective of this study is to generate chromosome-level assemblies of three avian species (saker falcon, budgerigar, and ostrich) previously reported as karyotypically rearranged compared to most birds. We also test the hypothesis that the density of conserved non-coding elements is associated with the positions of evolutionary breakpoint regions. 
Type Of Material Database/Collection of data 
Year Produced 2019 
Provided To Others? Yes  
URL https://datadryad.org/stash/dataset/doi:10.5061/dryad.q70q40m
 
Title Supplementary Material for: Fourth Report on Chicken Genes and Chromosomes 2022 
Description none 
Type Of Material Database/Collection of data 
Year Produced 2023 
Provided To Others? Yes  
URL https://karger.figshare.com/articles/dataset/Supplementary_Material_for_Fourth_Report_on_Chicken_Gen...
 
Title Supplementary Material for: Fourth Report on Chicken Genes and Chromosomes 2022 
Description none 
Type Of Material Database/Collection of data 
Year Produced 2023 
Provided To Others? Yes  
URL https://karger.figshare.com/articles/dataset/Supplementary_Material_for_Fourth_Report_on_Chicken_Gen...
 
Description List of collaborators 
Organisation Central Veterinary Research Laboratory
Country United Arab Emirates 
Sector Public 
PI Contribution A number of collaborations formed part of this project (see list)
Start Year 2006
 
Description List of collaborators 
Organisation Central Veterinary Research Laboratory
Country United Arab Emirates 
Sector Public 
PI Contribution A number of collaborations formed part of this project (see list)
Start Year 2006
 
Description List of collaborators 
Organisation Digital Scientific UK
Country United Kingdom 
Sector Private 
PI Contribution A number of collaborations formed part of this project (see list)
Start Year 2006
 
Description Multiple press coverage for work on avian genome evolution 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Our work led to the discovery of the likely dinosaur karyotype - this had extensive media coverage, including the BBC (see URL below)
Year(s) Of Engagement Activity 2018
URL https://www.bbc.co.uk/news/science-environment-44711974