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Mapping Genotypes to Phenotypes: Development of a Linkage Map and Mapping of Colour Polymorphisms in Ischnura elegans (Odonata)

Final Report Summary - COLOURFUL GENES (Mapping Genotypes to phenotypes: development of a linkage map and mapping of colour polymorphisms in Ischnura elegans (Odonata))

A summary description of the project objectives

Understanding the genetic basis of phenotypic variation remains one of the major challenges in biology. Linkage mapping provides means to identify genes that underlie ecologically important traits, such as colour polymorphisms, and this can be used to study the fitness consequences of variation at these loci. Genetic linkage maps are expected to become increasingly applied to ecological model organisms given their important role in linking phenotype to genotype, and the possibilities for using the results to address unanswered evolutionary and ecological questions.

In the proposed project, my objectives are:

(a) to construct the first linkage map for an Odonate insect, I. elegans;
(b) to map the female-limited colour polymorphism in the species and develop a specific marker that can be used to genotype males and larvae in which the colour polymorphisms is not expressed; and finally
(c) to use the novel microsatellites and the linkage map to evaluate the importance of gene flow and dispersal in the maintenance of this polymorphism in natural populations of I. elegans.

In line with the objectives of the Marie Curie actions, I will get advanced scientific training in molecular genomics which will greatly increase my research value and career perspectives. With the combination of novel molecular technology and ecological research, my project will contribute to enhance European Union (EU)'s scientific excellence in evolutionary biology and molecular genetics. Following the objectives of the work programme, my project links genotypes to phenotypes and will therefore provide the potential to identify and study the genetic basis of those traits affecting fitness that are key to natural selection in I. elegans. The research collaborations with people in Sweden, the Netherlands and Spain will provide opportunities to exchange ideas with several researchers active within my research field and initiate long-term international collaborations.

Description of the work performed since the beginning of the project

Objectives:

(1a) Development of microsatellites:
Several species specific microsatellites have been developed and tested, and have proven powerful in detecting gene flow and introgression in subsequent studies.

(1b) Genotyping of pedigree with microsatellites and amplified fragment length polymorphisms (AFLPs):
The pedigree has been genotyped with the microsatellites that we have developed, and we have improved the within pedigree group designations with this information. A large number of additional markers developed by restriction-site associated DNA (RAD) sequencing has also been performed for a large part of the individuals in the pedigree and are now being analysed.

(1c) Linkage mapping analyses:
First mapping analyses have been carried out with the microsatellites, and linked markers identified and sex specific recombination rates estimated.

(2a) Mapping of colour locus - development of genetic marker:
We use two approaches for this work. First, we conduct traditional linkage mapping using markers (RAD sequencing) and phenotyped individuals in a pedigree. Secondly, we use a genome-wide association (GWA) approach where we look for allele frequency differences between different groups of individuals, where each group correspond to one of the three focal female colour morphs. In the last analysis, we use whole genome sequencing of pools of several individuals.

(2b) Genotyping single nucleotide polymorphisms (SNPs) and comparative analyses:
Initial SNP analyses have been conducted and the comparative genome analyses have been initiated (a publication is in preparation).

(3) Analyse natural populations, estimate gene flow and morph related dispersal:
Natural populations have been analyses in detail. Gene flow among natural population is also a key focus.

Description of the main results achieved so far

Microsatellite development was carried out, and has since been used as tools to get a first linkage map in odonates. Although preliminary, this linkage map has already indicates sex biased recombination rates in this species. In Ischnura elegans, males are carry a X0 and females XX, which is identical to the model organisms Drosophila. In Drosophila, males show no recombination. In contrast, our linkage map suggests that males of I. elegans have a recombination rate, but that the rate of females is still estimates to be twice as high. The microsatellites have also been used to start population and landscape genetic studies, including a study to investigate hybridsation between I. elegans and its sister species in southern Europe. Several species specific microsatellites have been developed and tested, and have proven powerful in detecting gene flow and introgression in subsequent studies. We have genotyped individuals in a pedigree with microsatellites, and we have improved the within pedigree group designations with this information. We have genotyped a large part of the individuals in the pedigreea at a high number of additional markers by using the RAD sequencing technique, and this data is now being analysed. We have created the first linkage map in for Odonates, thereby linking markers into linkage groups and we detected sex specific recombination rates. We have been using two approaches for mapping the colour locus. First, we have conducting traditional linkage mapping using markers (RAD sequencing) and phenotyped individuals in a pedigree. Secondly, we use a GWA approach where we look for allele frequency differences between different groups of individuals, where each group correspond to one of the three focal female colour morphs. In the last analysis, we use whole genome sequencing of pools of several individuals.

Genotyping SNPs and comparative analyses: Initial SNP analyses have been conducted and the comparative genome analyses have been initiated (a publication is in preparation). Natural populations have been analysed in detail, and gene flow among natural population was also a key focus.

Expected final results and their potential impact and use (including the socio-economic impact and the wider societal implications of the project so far)

I have received training in molecular techniques, such as the development and application of microsatellite markers and population genetic and genomics. Training of software applications was successful (e.g. Geneious, Structure, and Geneland), and this knowledge has been transferred to other researchers at Lund University through teaching activities carried out by the researcher. Skills in writing research publications for the upper tier of journals in the field has been carried out during the project, as shown in the publication of several publication is renowned journal. Integration into the department and Sweden has been smooth, and the applicant has been involved in department and university wide groups, teaching and planning activities.