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Sunday, September 22, 2019

Reconstitution of eukaryotic chromosomes and manipulation of DNA N6-methyladenine alters chromatin and gene expression

DNA N6-adenine methylation (6mA) has recently been reported in diverse eukaryotes, spanning unicellular organisms to metazoans. Yet the functional significance of 6mA remains elusive due to its low abundance, difficulty of manipulation within native DNA, and lack of understanding of eukaryotic 6mA writers. Here, we report a novel DNA 6mA methyltransferase in ciliates, termed MTA1. The enzyme contains an MT-A70 domain but is phylogenetically distinct from all known RNA and DNA methyltransferases. Disruption of MTA1 in vivo leads to the genome-wide loss of 6mA in asexually growing cells and abolishment of the consensus ApT dimethylated motif. Genes exhibit subtle changes…

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Sunday, September 22, 2019

Insights into the biology of acidophilic members of the Acidiferrobacteraceae family derived from comparative genomic analyses.

The family Acidiferrobacteraceae (order Acidiferrobacterales) currently contains Gram negative, neutrophilic sulfur oxidizers such as Sulfuricaulis and Sulfurifustis, as well as acidophilic iron and sulfur oxidizers belonging to the Acidiferrobacter genus. The diversity and taxonomy of the genus Acidiferrobacter has remained poorly explored. Although several metagenome and bioleaching studies have identified its presence worldwide, only two strains, namely Acidiferrobacter thiooxydans DSM 2932T, and Acidiferrobacter spp. SP3/III have been isolated and made publically available. Using 16S rRNA sequence data publically available for the Acidiferrobacteraceae, we herein shed light into the molecular taxonomy of this family. Results obtained support the presence of three…

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Sunday, September 22, 2019

Improved nucleic acid extraction protocols for Ganoderma boninense, G. miniatocinctum and G. tornatum.

The first and most crucial step of all molecular techniques is to isolate high quality and intact nucleic acids. However, DNA and RNA isolation from fungal samples are usually difficult due to the cell walls that are relatively unsusceptible to lysis and often resistant to traditional extraction procedures. Although there are many extraction protocols for Ganoderma species, different extraction protocols have been applied to different species to obtain high yields of good quality nucleic acids, especially for genome and transcriptome sequencing. Ganoderma species, mainly G. boninense causes the basal stem rot disease, a devastating disease that plagues the oil palm…

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Sunday, September 22, 2019

Genome-scale analysis of Acetobacterium bakii reveals the cold adaptation of psychrotolerant acetogens by post-transcriptional regulation.

Acetogens synthesize acetyl-CoA via CO2 or CO fixation, producing organic compounds. Despite their ecological and industrial importance, their transcriptional and post-transcriptional regulation has not been systematically studied. With completion of the genome sequence of Acetobacterium bakii (4.28-Mb), we measured changes in the transcriptome of this psychrotolerant acetogen in response to temperature variations under autotrophic and heterotrophic growth conditions. Unexpectedly, acetogenesis genes were highly up-regulated at low temperatures under heterotrophic, as well as autotrophic, growth conditions. To mechanistically understand the transcriptional regulation of acetogenesis genes via changes in RNA secondary structures of 5′-untranslated regions (5′-UTR), the primary transcriptome was experimentally determined,…

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Sunday, September 22, 2019

Whole-genome landscape of Medicago truncatula symbiotic genes.

Advances in deciphering the functional architecture of eukaryotic genomes have been facilitated by recent breakthroughs in sequencing technologies, enabling a more comprehensive representation of genes and repeat elements in genome sequence assemblies, as well as more sensitive and tissue-specific analyses of gene expression. Here we show that PacBio sequencing has led to a substantially improved genome assembly of Medicago truncatula A17, a legume model species notable for endosymbiosis studies1, and has enabled the identification of genome rearrangements between genotypes at a near-base-pair resolution. Annotation of the new M. truncatula genome sequence has allowed for a thorough analysis of transposable elements…

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Sunday, September 22, 2019

Density-dependent enhanced replication of a densovirus in Wolbachia-infected Aedes cells is associated with production of piRNAs and higher virus-derived siRNAs.

The endosymbiotic bacterium Wolbachia pipientis has been shown to restrict a range of RNA viruses in Drosophila melanogaster and transinfected dengue mosquito, Aedes aegypti. Here, we show that Wolbachia infection enhances replication of Aedes albopictus densovirus (AalDNV-1), a single stranded DNA virus, in Aedes cell lines in a density-dependent manner. Analysis of previously produced small RNAs of Aag2 cells showed that Wolbachia-infected cells produced greater absolute abundance of virus-derived short interfering RNAs compared to uninfected cells. Additionally, we found production of virus-derived PIWI-like RNAs (vpiRNA) produced in response to AalDNV-1 infection. Nuclear fractions of Aag2 cells produced a primary vpiRNA…

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Sunday, September 22, 2019

Evolutionary conservation of Y Chromosome ampliconic gene families despite extensive structural variation.

Despite claims that the mammalian Y Chromosome is on a path to extinction, comparative sequence analysis of primate Y Chromosomes has shown the decay of the ancestral single-copy genes has all but ceased in this eutherian lineage. The suite of single-copy Y-linked genes is highly conserved among the majority of eutherian Y Chromosomes due to strong purifying selection to retain dosage-sensitive genes. In contrast, the ampliconic regions of the Y Chromosome, which contain testis-specific genes that encode the majority of the transcripts on eutherian Y Chromosomes, are rapidly evolving and are thought to undergo species-specific turnover. However, ampliconic genes are…

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Sunday, September 22, 2019

Genomic and transcriptomic comparisons of closely related malaria parasites differing in virulence and sequestration pattern.

Background: Malaria parasite species differ greatly in the harm they do to humans. While P. falciparum kills hundreds of thousands per year, P. vivax kills much less often and P. malariae is relatively benign. Strains of the rodent malaria parasite Plasmodium chabaudi show phenotypic variation in virulence during infections of laboratory mice. This make it an excellent species to study genes which may be responsible for this trait. By understanding the mechanisms which underlie differences in virulence we can learn how parasites adapt to their hosts and how we might prevent disease. Methods: Here we present a complete reference genome…

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Sunday, September 22, 2019

Transcriptional landscape of a blaKPC-2 plasmid and response to imipenem exposure in Escherichia coli TOP10.

The diffusion of KPC-2 carbapenemase is closely related to the spread of Klebsiella pneumoniae of the clonal-group 258 and linked to IncFIIK plasmids. Little is known about the biology of multi-drug resistant plasmids and the reasons of their successful dissemination. Using E. coli TOP10 strain harboring a multi-replicon IncFIIK-IncFIB blaKPC-2-gene carrying plasmid pBIC1a from K. pneumoniae ST-258 clinical isolate BIC-1, we aimed to identify basal gene expression and the effects of imipenem exposure using whole transcriptome approach by RNA sequencing (RNA-Seq). Independently of the antibiotic pressure, most of the plasmid-backbone genes were expressed at low levels. The most expressed pBIC1a…

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Saturday, September 21, 2019

PacBio assembly of a Plasmodium knowlesi genome sequence with Hi-C correction and manual annotation of the SICAvar gene family.

Plasmodium knowlesi has risen in importance as a zoonotic parasite that has been causing regular episodes of malaria throughout South East Asia. The P. knowlesi genome sequence generated in 2008 highlighted and confirmed many similarities and differences in Plasmodium species, including a global view of several multigene families, such as the large SICAvar multigene family encoding the variant antigens known as the schizont-infected cell agglutination proteins. However, repetitive DNA sequences are the bane of any genome project, and this and other Plasmodium genome projects have not been immune to the gaps, rearrangements and other pitfalls created by these genomic features.…

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Saturday, September 21, 2019

A flexible and efficient template format for circular consensus sequencing and SNP detection.

A novel template design for single-molecule sequencing is introduced, a structure we refer to as a SMRTbell template. This structure consists of a double-stranded portion, containing the insert of interest, and a single-stranded hairpin loop on either end, which provides a site for primer binding. Structurally, this format resembles a linear double-stranded molecule, and yet it is topologically circular. When placed into a single-molecule sequencing reaction, the SMRTbell template format enables a consensus sequence to be obtained from multiple passes on a single molecule. Furthermore, this consensus sequence is obtained from both the sense and antisense strands of the insert…

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