In this study, we applied the iterative procedure (IP) method to search for families of highly diverged dispersed repeats in the genome of Cyanidioschyzon merolae, which contains over 16 million bases. The algorithm included the construction of position weight matrices (PWMs) for repeat families and the identification of more dispersed repeats based on the PWMs using dynamic programming. The results showed that the C. merolae genome contained 20 repeat families comprising a total of 33,938 dispersed repeats, which is significantly more than has been previously found using other methods. The repeats varied in length from 108 to 600 bp (522.54 bp in average) and occupied more than 72% of the C. merolae genome, whereas previously identified repeats, including tandem repeats, have been shown to constitute only about 28%. The high genomic content of dispersed repeats and their location in the coding regions suggest a significant role in the regulation of the functional activity of the genome.

Sex chromosomes have evolved in many plant species with separate sexes. Current plant research is shifting from examining the structure of sex chromosomes to exploring their functional aspects. New studies are progressively unveiling the specific genetic and epigenetic mechanisms responsible for shaping distinct sexes in plants. While the fundamental methods of molecular biology and genomics are generally employed for the analysis of sex chromosomes, it is often necessary to modify classical procedures not only to simplify and expedite analyses but sometimes to make them possible at all. In this review, we demonstrate how, at the level of structural and functional genetics, cytogenetics, and bioinformatics, it is essential to adapt established procedures for sex chromosome analysis.

Polyploidy is a widespread phenomenon across angiosperms, and one of the main drivers of diversification. Whilst it frequently involves hybridisation, autopolyploidy is also an important feature of plant evolution. Minority cytotypes are frequently overlooked due to their lower frequency in populations, but the development of techniques such as flow cytometry, which enable the rapid screening of cytotype diversity across large numbers of individuals, is now providing a more comprehensive understanding of cytotype diversity within species. Senecio doronicum is a relatively common daisy found throughout European mountain grasslands from subalpine to almost nival elevations. We have carried out a population-level cytotype screening of 500 individuals from Tête Grosse (Alpes-de-Haute-Provence, France), confirming the coexistence of tetraploid (28.2%) and octoploid cytotypes (71.2%), but also uncovering a small number of hexaploid individuals (0.6%). The analysis of repetitive elements from short-read genome-skimming data combined with nuclear (ITS) and whole plastid DNA sequences support an autopolyploid origin of the polyploid S. doronicum individuals and provide molecular evidence regarding the sole contribution of tetraploids in the formation of hexaploid individuals. The evolutionary impact and resilience of the new cytotype have yet to be determined, although the coexistence of different cytotypes may indicate nascent speciation.

Sequencing the giga-genomes of several pine species has enabled comparative genomic analyses of these outcrossing tree species. Previous studies have revealed the wide distribution and extraordinary diversity of transposable elements (TEs) that occupy the large intergenic spaces in conifer genomes. In this study, we analyzed the distribution of TEs in gene regions of the assembled genomes of Pinus taeda and Pinus lambertiana using high-performance computing resources. The quality of draft genomes and the genome annotation have significant consequences for the investigation of TEs and these aspects are discussed. Several TE families frequently inserted into genes or their flanks were identified in both species\' genomes. Potentially important sequence motifs were identified in TEs that could bind additional regulatory factors, promoting gene network formation with faster or enhanced transcription initiation. Node genes that contain many TEs were observed in multiple potential transposable element-associated networks. This study demonstrated the increased accumulation of TEs in the introns of stress-responsive genes of pines and suggests the possibility of rewiring them into responsive networks and sub-networks interconnected with node genes containing multiple TEs. Many such regulatory influences could lead to the adaptive environmental response clines that are characteristic of naturally spread pine populations.

UNASSIGNED: Nesting is common in LTR retrotransposons, especially in large genomes containing a high number of elements.
UNASSIGNED: We analyzed 12 plant genomes and obtained 1491 pairs of nested and original (pre-existing) LTR retrotransposons. We systematically analyzed mutual nesting of individual LTR retrotransposons and found that certain families, more often belonging to the Ty3/gypsy than Ty1/copia superfamilies, showed a higher nesting frequency as well as a higher preference for older copies of the same family (\"autoinsertions\"). Nested LTR retrotransposons were preferentially located in the 3\'UTR of other LTR retrotransposons, while coding and regulatory regions (LTRs) are not commonly targeted. Insertions displayed a weak preference for palindromes and were associated with a strong positional pattern of higher predicted nucleosome occupancy. Deviation from randomness in target site choice was also found in 13,983 non-nested plant LTR retrotransposons.
UNASSIGNED: We reveal that nesting of LTR retrotransposons is not random. Integration is correlated with sequence composition, secondary structure and the chromatin environment. Insertion into retrotransposon positions with a low negative impact on family fitness supports the concept of the genome being viewed as an ecosystem of various elements.

UNASSIGNED: Kinetoplastids are a flagellated group of protists, including some parasites, such as Trypanosoma and Leishmania species, that can cause diseases in humans and other animals. The genomes of these species enclose a fraction of retrotransposons including VIPER and TATE, two poorly studied transposable elements that encode a tyrosine recombinase (YR) and were previously classified as DIRS elements. This study investigated the distribution and evolution of VIPER and TATE in kinetoplastids to understand the relationships of these elements with other retrotransposons.
UNASSIGNED: We observed that VIPER and TATE have a discontinuous distribution among Trypanosomatidae, with several events of loss and degeneration occurring during a vertical transfer evolution. We were able to identify the terminal repeats of these elements for the first time, and we showed that these elements are potentially active in some species, including T. cruzi copies of VIPER. We found that VIPER and TATE are strictly related elements, which were named in this study as VIPER-like. The reverse transcriptase (RT) tree presented a low resolution, and the origin and relationships among YR groups remain uncertain. Conversely, for RH, VIPER-like grouped with Hepadnavirus, whereas for YR, VIPER-like sequences constituted two different clades that are closely allied to Crypton. Distinct topologies among RT, RH and YR trees suggest ancient rearrangements/exchanges in domains and a modular pattern of evolution with putative independent origins for each ORF.
UNASSIGNED: Due to the presence of both elements in Bodo saltans, a nontrypanosomatid species, we suggested that VIPER and TATE have survived and remained active for more than 400 million years or were reactivated during the evolution of the host species. We did not find clear evidence of independent origins of VIPER-like from the other YR retroelements, supporting the maintenance of the DIRS group of retrotransposons. Nevertheless, according to phylogenetic findings and sequence structure obtained by this study and other works, we proposed separating DIRS elements into four subgroups: DIRS-like, PAT-like, Ngaro-like, and VIPER-like.

Transposable elements (TEs) are ubiquitous in eukaryotic genomes and their mobility impacts genome structure and function in myriad ways. Because of their abundance, activity, and repetitive nature, the characterization and analysis of TEs remain challenging, particularly from short-read sequencing projects. To overcome this difficulty, we have developed a method that estimates TE copy number from short-read sequences. To test the accuracy of our method, we first performed an in silico analysis of the reference Sorghum bicolor genome, using both reference-based and de novo approaches. The resulting TE copy number estimates were strikingly similar to the annotated numbers. We then tested our method on real short-read data by estimating TE copy numbers in several accessions of S. bicolor and its close relative S. propinquum. Both methods effectively identify and rank similar TE families from highest to lowest abundance. We found that de novo characterization was effective at capturing qualitative variation, but underestimated the abundance of some TE families, specifically families of more ancient origin. Also, interspecific reference-based mapping of S. propinquum reads to the S. bicolor database failed to fully describe TE content in S. propinquum, indicative of recent TE activity leading to changes in the respective repetitive landscapes over very short evolutionary timescales. We conclude that reference-based analyses are best suited for within-species comparisons, while de novo approaches are more reliable for evolutionarily distant comparisons.

Microsatellite DNA families (MDF) are stretches of DNA that share similar or identical sequences beside nuclear simple-sequence repeat (nSSR) motifs, potentially causing problems during nSSR marker development. Primers positioned within MDFs can bind several times within the genome and might result in multiple banding patterns. It is therefore common practice to exclude MDF loci in the course of marker development. Here, we propose an approach to deal with multiple primer-binding sites by purposefully positioning primers within the detected repetitive element. We developed a new protocol to determine the family type and the primer position in relation to MDFs using the software packages repark and repeatmasker together with an in-house R script. We re-evaluated newly developed nSSR markers for the lepidopteran Marbled White (Melanargia galathea) and explored the implications of our results with regard to published data sets of the butterfly Euphydryas aurinia, the grasshopper Stethophyma grossum, the conifer Pinus cembra and the crucifer Arabis alpina. For M. galathea, we show that it is not only possible to develop reliable nSSR markers for MDF loci, but even to benefit from their presence in some cases: We used one unlabelled primer, successfully binding within an MDF, for two different loci in a multiplex PCR, combining this family primer with uniquely binding and fluorescently labelled primers outside of MDFs, respectively. As MDFs are abundant in many taxa, we propose to consider these during nSSR marker development in taxa concerned. Our new approach might help in reducing the number of tested primers during nSSR marker development.

BACKGROUND: As Short Interspersed Elements (SINEs), human-specific Alu elements can be used for population genetic studies. Very recent inserts are polymorphic within and between human populations. In a sample of 30 elements originating from three different Alu subfamilies, we investigated whether they are preserved in prehistorical skeletal human remains from the Bronze Age Lichtenstein cave in Lower Saxony, Germany. In the present study, we examined a prehistoric triad of father, mother and daughter.
RESULTS: For 26 of the 30 Alu loci investigated, definite results were obtained. We were able to demonstrate that presence/absence analyses of Alu elements can be conducted on individuals who lived 3,000 years ago. The preservation of the ancient DNA (aDNA) is good enough in two out of three ancient individuals to routinely allow the amplification of 500 bp fragments. The third individual revealed less well-preserved DNA, which results in allelic dropout or complete amplification failures. We here present an alternative molecular approach to deal with these degradation phenomena by using internal Alu subfamily specific primers producing short fragments of approximately 150 bp.
CONCLUSIONS: Our data clearly show the possibility of presence/absence analyses of Alu elements in individuals from the Lichtenstein cave. Thus, we demonstrate that our method is reliably applicable for aDNA samples with good or moderate DNA preservation. This method will be very useful for further investigations with more Alu loci and larger datasets. Human population genetic studies and other large-scale investigations would provide insight into Alu SINE-based microevolutionary processes in humans during the last few thousand years and help us comprehend the evolutionary dynamics of our genome.

Evidence continues to grow on potential environmental health hazards associated with engineered nanomaterials (ENMs). While the geno- and cytotoxic effects of ENMs have been investigated, their potential to target the epigenome remains largely unknown. The aim of this study is two-fold: 1) determining whether or not industry relevant ENMs can affect the epigenome in vivo and 2) validating a recently developed in vitro epigenetic screening platform for inhaled ENMs. Laser printer-emitted engineered nanoparticles (PEPs) released from nano-enabled toners during consumer use and copper oxide (CuO) were chosen since these particles induced significant epigenetic changes in a recent in vitro companion study. In this study, the epigenetic alterations in lung tissue, alveolar macrophages and peripheral blood from intratracheally instilled mice were evaluated. The methylation of global DNA and transposable elements (TEs), the expression of the DNA methylation machinery and TEs, in addition to general toxicological effects in the lung were assessed. CuO exhibited higher cell-damaging potential to the lung, while PEPs showed a greater ability to target the epigenome. Alterations in the methylation status of global DNA and TEs, and expression of TEs and DNA machinery in mouse lung were observed after exposure to CuO and PEPs. Additionally, epigenetic changes were detected in the peripheral blood after PEPs exposure. Altogether, CuO and PEPs can induce epigenetic alterations in a mouse experimental model, which in turn confirms that the recently developed in vitro epigenetic platform using macrophage and epithelial cell lines can be successfully utilized in the epigenetic screening of ENMs.