Diving deep into digital literacy:emerging methods for research

Literacy studies approaches have tended to adopt a position which enables ethnographic explorations of a wide range of ‘literacies’. An important issue arising is the new challenge required for researchers to capture, manage, and analyse data that highlight the unique character of practices around texts in digital environments. Such inquiries, we argue, require multiple elements of data to be captured and analysed as part of effective literacy ethnographies. These include such things as the unfolding of digital texts, the activities around them, and features of the surrounding social and material environment. This paper addresses these methodological issues drawing from three educationally focused studies, and reporting their experiences and insights within uniquely different contexts. We deal with the issue of adopting new digital methods for literacy research through the notion of a ‘deep dive’ to explore educational tasks in classrooms. Through a discussion of how we approached the capture and analysis of our data, we present methods to better understand digital literacies in education. We then outline challenges posed by our methods, how they can be used more broadly for researching interaction in digital environments, and how they augment transdisciplinary debates and trends in research methods

Reprogrammed Transcriptome in Rhesus-Bovine Interspecies Somatic Cell Nuclear Transfer Embryos

Global activation of the embryonic genome (EGA), one of the most critical steps in early mammalian embryo development, is recognized as the time when interspecies somatic cell nuclear transfer (iSCNT) embryos fail to thrive.In this study, we analyzed the EGA-related transcriptome of rhesus-bovine iSCNT 8- to 16-cell embryos and dissected the reprogramming process in terms of embryonic gene activation, somatic gene silencing, and maternal RNA degradation. Compared with fibroblast donor cells, two thousand and seven genes were activated in iSCNT embryos, one quarter of them reaching expression levels comparable to those found in in vitro fertilized (IVF) rhesus embryos. This suggested that EGA in iSCNT embryos had partially recapitulated rhesus embryonic development. Eight hundred and sixty somatic genes were not silenced properly and continued to be expressed in iSCNT embryos, which indicated incomplete nuclear reprogramming. We compared maternal RNA degradation in bovine oocytes between bovine-bovine SCNT and iSCNT embryos. While maternal RNA degradation occurred in both SCNT and iSCNT embryos, we saw more limited overall degradation of maternal RNA in iSCNT embryos than in SCNT embryos. Several important maternal RNAs, like GPF9, were not properly processed in SCNT embryos.Our data suggested that iSCNT embryos are capable of triggering EGA, while a portion of somatic cell-associated genes maintain their expression. Maternal RNA degradation seems to be impaired in iSCNT embryos. Further understanding of the biological roles of these genes, networks, and pathways revealed by iSCNT may expand our knowledge about cell reprogramming, pluripotency, and differentiation

Genome-Wide Analysis of GLD-1–Mediated mRNA Regulation Suggests a Role in mRNA Storage

Translational repression is often accompanied by mRNA degradation. In contrast, many mRNAs in germ cells and neurons are “stored" in the cytoplasm in a repressed but stable form. Unlike repression, the stabilization of these mRNAs is surprisingly little understood. A key player in Caenorhabditis elegans germ cell development is the STAR domain protein GLD-1. By genome-wide analysis of mRNA regulation in the germ line, we observed that GLD-1 has a widespread role in repressing translation but, importantly, also in stabilizing a sub-population of its mRNA targets. Additionally, these mRNAs appear to be stabilized by the DDX6-like RNA helicase CGH-1, which is a conserved component of germ granules and processing bodies. Because many GLD-1 and CGH-1 stabilized mRNAs encode factors important for the oocyte-to-embryo transition (OET), our findings suggest that the regulation by GLD-1 and CGH-1 serves two purposes. Firstly, GLD-1–dependent repression prevents precocious translation of OET–promoting mRNAs. Secondly, GLD-1– and CGH-1–dependent stabilization ensures that these mRNAs are sufficiently abundant for robust translation when activated during OET. In the absence of this protective mechanism, the accumulation of OET–promoting mRNAs, and consequently the oocyte-to-embryo transition, might be compromised

Cloning and characterization of a novel oocyte-specific gene encoding an F-Box protein in rainbow trout (Oncorhynchus mykiss)

BACKGROUND: Oocyte-specific genes play critical roles in oogenesis, folliculogenesis and early embryonic development. The objectives of this study were to characterize the expression of a novel oocyte-specific gene encoding an F-box protein during ovarian development in rainbow trout, and identify its potential interacting partners in rainbow trout oocytes. METHODS: Through analysis of expressed sequence tags (ESTs) from a rainbow trout oocyte cDNA library, a novel transcript represented by ESTs only from the oocyte library was identified. The complete cDNA sequence for the novel gene (named fbxoo) was obtained by assembling sequences from an EST clone and a 5′RACE product. The expression and localization of fbxoo mRNA and protein in ovaries of different developmental stages were analyzed by quantitative real time PCR, immunoblotting, in situ hybridization and immunohistochemistry. Identification of Fbxoo binding proteins was performed by yeast two-hybrid screening. RESULTS: fbxoo mRNA is specifically expressed in mature oocytes as revealed by tissue distribution analysis. The fbxoo cDNA sequence is 1,996 bp in length containing an open reading frame, which encodes a predicted protein of 514 amino acids. The novel protein sequence does not match any known protein sequences in the NCBI database. However, a search of the Pfam protein database revealed that the protein contains an F-box motif at the N-terminus, indicating that Fbxoo is a new member of the F-box protein family. The expression of fbxoo mRNA and protein is high in ovaries at early pre-vitellogenesis stage, and both fbxoo mRNA and protein are predominantly expressed in early pre-vitellogenic oocytes. Several proteins including tissue inhibitor of metalloproteinase 2 (Timp2) were identified as potential Fbxoo protein binding partners. CONCLUSIONS: Results suggest that the novel oocyte-specific F-box protein may play an important role in early oocyte development by regulating other critical proteins involved in oogenesis in rainbow trout

Segments containing alternating purine and pyrimidine dinucleotides: patterns of polymorphism in humans and prevalence throughout phylogeny.

Tandem dinucleotide repeats of GT or AC [(GT)n/(AC)n] where n greater than or equal to 14 are highly polymorphic and other simple repeats such as (CT)n/(AG)n and (A)n(T)n are also polymorphic. The uniformity of these sequences precludes a mechanistic differentiation between recombination or polymerase slippage. Since (GT)n/(AC)n or (CT)n/(AG)n segments of desired size were not available in our gene of interest, we analyzed a 187+ bp segment in the factor IX gene with multiple short dinucleotide repeats. This sequence contains a melody of short dinucleotide repeats which includes a 142+ bp segment of alternating purines and pyrimidines. Amplification of this sequence in 167 individuals of different ethnicity and direct sequencing of 106 individuals (23 kb of sequence) failed to reveal simple variation in the number of tandem dinucleotide repeats. However, polymorphism in the 142+ alternating purine and pyrimidine segment (RY)n was detected due to the insertion of two related repeat units of 24 bp (A) and 26 bp (B). Two previously described alleles (AB, A2B2) and two novel presumptive recombinants were found (A2B, A3B2) for a total of four alleles. An analysis of (RY)n segments in GenBank revealed an extraordinary enrichment in the genome of mammals, invertebrates, and yeast and a marked reduction in bacteria. Rodent (RY)n were larger and substantially more frequent than those in primates. When a second (RY)n was examined in the exon 8 of human factor IX gene, it was polymorphic at short repeats of (GT)n/(AC)n (n = 3-6) in Western Europeans and Koreans. In addition, an (RY)n in the dystrophin gene had four polymorphic alleles involving AT and GT dinucleotides. Thus (RY)n segments appear to be abundant and highly polymorphic. The asymmetric patterns of polymorphism and the absence of simple dinucleotide variation in 23 kb of sequence are compatible with recombination or sister chromatid exchange, but not polymerase slippage. By inference, recombination should underlie the polymorphisms at (GT)n/(AC)n since they are a subset of (RY)n and they commonly occur in the context of longer (RY)n

The Anatomy to Genomics (ATG) Start Genetics medical school initiative: incorporating exome sequencing data from cadavers used for Anatomy instruction into the first year curriculum

Abstract Background The increasing use of next generation DNA sequencing in clinical medicine is exposing the need for more genetics education in physician training. We piloted an initiative to determine the feasibility of incorporating exome sequencing data generated from DNA obtained from cadavers used for teaching Anatomy into a first year medical student integrated block-style course. Methods We optimized the procedure to obtain DNA for exome sequencing by comparing the quality and quantity of DNA isolated from several tissues by two different extraction methods. DNA was sequenced using exome capture and analyzed using standard methods. Single nucleotide variants (SNVs), as well as small insertions/deletions, with potential functional impact were selected by faculty for student teams to independently investigate and prepare presentations on their findings. Results A total of seven cadaver DNAs were sequenced yielding high quality results. SNVs were identified that were associated, with known physical traits and disease susceptibility, as well as pharmacogenomic phenotypes. Students presented findings based on correlation with known clinical information about the cadavers’ diseases and traits. Conclusion Exome sequencing of cadaver DNA is a useful tool to integrate Anatomy with Genetics and Biochemistry into a first year medical student core curriculum

Exome-wide analysis of copy number variation shows association of the human leukocyte antigen region with asthma in UK Biobank

Background: The role of copy number variants (CNVs) in susceptibility to asthma is not well understood. This is, in part, due to the difficulty of accurately measuring CNVs in large enough sample sizes to detect associations. The recent availability of whole-exome sequencing (WES) in large biobank studies provides an unprecedented opportunity to study the role of CNVs in asthma. Methods: We called common CNVs in 49,953 individuals in the first release of UK Biobank WES using ClinCNV software. CNVs were tested for association with asthma in a stage 1 analysis comprising 7098 asthma cases and 36,578 controls from the first release of sequencing data. Nominally-associated CNVs were then meta-analysed in stage 2 with an additional 17,280 asthma cases and 115,562 controls from the second release of UK Biobank exome sequencing, followed by validation and fine-mapping. Results: Five of 189 CNVs were associated with asthma in stage 2, including a deletion overlapping the HLA-DQA1 and HLA-DQB1 genes, a duplication of CHROMR/PRKRA, deletions within MUC22 and TAP2, and a duplication in FBRSL1. The HLA-DQA1, HLA-DQB1, MUC22 and TAP2 genes all reside within the human leukocyte antigen (HLA) region on chromosome 6. In silico analyses demonstrated that the deletion overlapping HLA-DQA1 and HLA-DQB1 is likely to be an artefact arising from under-mapping of reads from non-reference HLA haplotypes, and that the CHROMR/PRKRA and FBRSL1 duplications represent presence/absence of pseudogenes within the HLA region. Bayesian fine-mapping of the HLA region suggested that there are two independent asthma association signals. The variants with the largest posterior inclusion probability in the two credible sets were an amino acid change in HLA-DQB1 (glutamine to histidine at residue 253) and a multi-allelic amino acid change in HLA-DRB1 (presence/absence of serine, glycine or leucine at residue 11). Conclusions: At least two independent loci characterised by amino acid changes in the HLA-DQA1, HLA-DQB1 and HLA-DRB1 genes are likely to account for association of SNPs and CNVs in this region with asthma. The high divergence of haplotypes in the HLA can give rise to spurious CNVs, providing an important, cautionary tale for future large-scale analyses of sequencing data