Undertanding The Contribution Of Tandem Repeats Togenetic Variations In Bumblebee Genome

Bumblebees(Hymenoptera: Apidae)are a group of pollinating insects that belong to the genus Bombus,which play an important role in promoting agricultural production and maintaining the balance of natural ecosystem.Pollination services provided by bumblebees will not only improve the quality and yield of crops,but also reduce the use of chemical hormones,with obvious economic and ecological benefits.Because of the large scale of facility agriculture in China,the market potential of using bumblebee for pollination service is huge.However,despite the abundant species resources of bumblebees in China,there is no native bumblebee with excellent pollination and breeding performance,so bumblebees currently used in pollination services in China are mainly from importing,which has the risk of biological invasion.In order to cultivate native bumblebees with excellent traits for pollination service,the mining and utilization of important functional genes of bumblebee is one of the strategies to solve the problem.Tandem repeats are the hot spots of mutation in the genome.Their mutations are an important source of genetic variation.In addition,their mutations could promote variations in biological traits.In this study,the contribution of tandem repeats to the genetic variation of regulatory and coding regions of bumblebee genes was investigated,with obtaining the following results:1.The regulatory regions of bumblebee genes were identified.We conducted ATAC-seq experiments on four developmental stages of bumblebee.According to the information of the open chromatin regions obtained in the experiment,the regulatory region of bumblebee gene was identified by bioinformatics method.The annotation of gene regulatory sequences in bumblebees closed the otherwise gap.2.Tandem repeats promoted the genetic differentiation of inter-specific and intra-specific regulatory sequences of bumblebee genes.Based on the annotation information of tandem repeats and regulatory regions in B.terrestris genome,a total of 3121 tandem repeats were found within the regulatory sequences of bumblebee genes.Comparative genomics analysis between two species of subgenus Bombus revealed 185 polymorphic tandem repeats loci between the two species.That is,tandem repeats of these loci promoted the genetic differentiation of inter-specific regulatory sequences of bumblebee genes.In addition,a tandem repeat locus was found in the promoter region of LOC100645194,which exhibits polymorphic between the wild and domesticated B.terrestris.Functional classification of genes whose regulatory sequences were driven by tandem repeats suggested that they participate in the regulation of nitrogen metabolism,cell metabolism,cell communication,signal transmission and other important biological processes etc.3.Polyclonal antibodies that target proteins affected by tandem repeats were successfully prepared.We selected three genes with inter-specific tandem repeat variation in the protein-coding regions and one gene with intra-specific tandem repeat variation in protein-coding region of B.terrestris(wild and domesticated).Prokaryotic expression of recombinant proteins and preparation of polyclonal antibodies were performed on those four genes.Polyclonal antibodies with high quality,high potency and good specificity were obtained.The antibodies have laid a foundation for studying the biological function of the investigated bumblebee bees.Because the variation of tandem repeats in these genes leads to the variation in the length of the amino acid sequence encoded by this gene between and within bumblebee species,the antibodies prepared in this study are also useful to study the effect of mutations produced by tandem repeats on protein function.