The overall goal of this project is to investigate how natural selection maintains variation in immune genes, using a wild rodent (the bank vole) as study system. We have re-sequenced a population sample of bank voles to identify immune genes under balancing selection, i.e. natural selection that acts to maintain genetic variation. We have also performed analyses to investigate if factors like gene expression patterns determine which genes are under balancing selection, and the functional effects of polymorphisms in immune genes under balancing selection; these studies are now nearing completion. During 2025, we aim to address two new questions: (i) To what extent is balancing selection population-specific? (ii) What is the relative importance of balancing selection within populations and divergent selection among populations as drivers of global genetic variation? For this purpose, we will resequence bank voles from additional populations. Our study species, the bank vole (Myodes glareolus), is widely distributed in Europe and is a popular study species for various ecological and evolutionary questions. Råberg’s group has been using the bank vole as study species for research on the evolutionary ecology of host-pathogen interactions since 2006. In 2017 we performed whole-genome resequencing at 40× using Illumina HiSeq X of 30 individual bank voles from our study population in southern Sweden. This data has been used for a genome-wide scan for signatures of balancing selection as well as analyses of selective constraint on protein coding sequences. In 2021, we performed in vitro assays of splenocyte responses to bacteria by RNA-sequencing. All in all, 169 samples from 63 bank voles were sequenced. In 2023, we performed RNA-sequencing of 10 tissues from 2-5 males and 2-5 females from both summer and winter, to measure level and breadth of gene expression (all in all 95 samples). During 2024 we sequenced the genome of a grey-sided vole (a closely related species to the bank vole); we use this as an outgroup to enhance the power of selection analyses in the bank vole. During 2025 we plan to re-sequence a population sample of bank voles (c. 25) from northern Sweden, to test for population differentiation and population-specific signatures of balancing selection, as outlined above (DNA extraction is performed currently). The storage project will be used for analyses of all these data sets, but primarily the data sets with resequencing data.