Genome-wide determinants of Tn5 insertion preference
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus Homo sapiens
PROVIDER: GSE164997 | GEO | 2021/10/01
REPOSITORIES: GEO
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Project description:Genome-wide determinants of Tn5 insertion preference
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2014-07-14 | GSE58652 | GEO
Project description:Abstract Tn5 transposase, which can efficiently tagment the genome, has been widely adopted as a molecular tool in next-generation sequencing, from short-read sequencing to more complex methods such as assay for transposase-accessible chromatin using sequencing (ATAC-seq). Here, we systematically map Tn5 insertion characteristics across several model organisms, finding critical parameters that affect its insertion. On naked genomic DNA, we found that Tn5 insertion is not uniformly distributed or random. To uncover drivers of these biases, we used a machine learning framework, which revealed that DNA shape cooperatively works with DNA motif to affect Tn5 insertion preference. These intrinsic insertion preferences can be modeled using nucleotide dependence information from DNA sequences, and we developed a computational pipeline to correct for these biases in ATAC-seq data. Using our pipeline, we show that bias correction improves the overall performance of ATAC-seq peak detection, recovering many potential false-negative peaks. Furthermore, we found that these peaks are bound by transcription factors, underscoring the biological relevance of capturing this additional information. These findings highlight the benefits of an improved understanding and precise correction of Tn5 insertion preference.
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Project description:Tn5 transposase is used as a tool for detecting nucleosome-free regions of genomic DNA in eukaryotes, but its DNA target site in chromatin has not been understood. In the present study, the well-positioned dinucleosomes were reconstituted, and the Tn5 transposase target sites were mapped in the dinucleosomes in vitro. We found that Tn5 transposase preferentially targets near the entry-exit DNA region within the nucleosome, if the linker DNA exists between two nucleosomes. This specific DNA targeting by Tn5 did not depend on the linker DNA length and DNA sequence. Tn5 transposase becomes to target the middle of the linker DNA, in addition to the entry-exit site of the nucleosome, if the linker DNA length extends to 30 base pairs. These in vitro data provide direct evidence for the Tn5 target sites in the nucleosome, resulting important information for interpretation of the Tn5-transposase-based genomics methods, which have been interpreted as linker or nucleosome-free DNA regions in genomes.
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Project description:Tn5 transposon tagments double-strand DNA and RNA/DNA hybrid to generate nucleic acids ready to be amplified for high throughput sequencing. The nucleic acid substrates of Tn5 transposon need to be explored to increase the applications of Tn5. Here, we find that Tn5 transposon is able to transport oligos into the 5’ end of single-strand DNA, which are more than 140 base pairs of nucleic acids. Based on this property of Tn5, we develop a tagmentation-based and ligation-enabled single-strand DNA sequencing method, TABLE-seq. Through a series of reaction temperature, time, and enzyme concentration tests, we managed to apply this TABLE-seq to strand-specific RNA sequencing, starting with as low as 30 picograms of total RNA. Moreover, compared with traditional dUTP based strand-specific RNA sequencing, this method detects more genes, has higher strand-specificity, and shows more evenly distributed reads across genes. Together, our results provide insights into the properties of Tn5 transposons and expand the applications of Tn5 in cutting-edge sequencing techniques.
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Project description:Social interactions can drive distinct gene expression profiles which may vary by social context. Here we use female sailfin molly fish (Poecilia latipinna) to identify genomic profiles associated with preference behavior in distinct social contexts: male-interactions (mate choice) versus female-interactions (shoaling partner preference). We measured behavior of 15 females interacting in a non-contact environment with either two males or two females for 30 minutes followed by whole brain transcriptomic profiling by RNA sequencing. We profiled females that exhibited high levels of social affiliation and great variation in preference behavior to identify an order of magnitude more differentially expressed genes associated with behavioral variation than by differences in social context. Using linear modeling (limma), we took advantage of the individual variation in preference behavior to identify unique gene sets that exhibited distinct correlational patterns of expression with preference behavior in each social context. By combining limma and weighted gene co-expression network analyses (WGCNA) approaches we identify a refined set of 401 genes robustly associated with mate preference that is independent of shoaling partner preference or general social affiliation. While our refined gene set confirmed neural plasticity pathways involved in moderating female preference behavior, we also identified a significant proportion of discovered that our preference-associated genes were enriched for ‘immune system’ gene ontology categories. We hypothesize that the association between mate preference and transcriptomic immune function is driven by the less well-known role of these genes in neural plasticity which is likely involved in higher-order learning and processing during mate choice decisions.
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