Building a reference transcriptome for Juniperus squamata (Cupressaceae) based on single-molecule real-time sequencing

Objectives Cupressaceae is the second largest family of coniferous trees (Coniferopsida) with important economic and ecological values. However, like other conifers, the members of Cupressaceae have extremely large genome (> 8 gigabytes), which limited the researches of these taxa. A high-quality transcriptome is an important resource for gene discovery and annotation for non-model organisms. Data description Juniperus squamata, a tetraploid species which is widely distributed in Asian mountains, represents the largest genus, Juniperus, in Cupressaceae. Single-molecule real-time sequencing was used to obtain full-length transcriptome of Juniperus squamata. The full-length transcriptome was corrected with Illumina RNA-seq data from the same individual. A total of 47,860 non-redundant full-length transcripts, N50 of which was 2839, were obtained. A total of 57,393 simple sequence repeats were identified and 268,854 open reading frames were predicted for Juniperus squamata. A BLAST alignment against non-redundant protein database was conducted and 10,818 sequences were annotated in Gene Ontology database. InterPro analysis shows that 30,403 sequences have been functionally characterized against its member database. This data presents the first comprehensive transcriptome characterization of Juniperus species, and provides an important reference for researches on the genomics and evolutionary history of Cupressaceae plants and conifers in the future.

Taxonomy-aware, sequence similarity ranking reliably predicts phage–host relationships

Background Characterizing phage–host interactions is critical to understanding the ecological role of both partners and effective isolation of phage therapeuticals. Unfortunately, experimental methods for studying these interactions are markedly slow, low-throughput, and unsuitable for phages or hosts difficult to maintain in laboratory conditions. Therefore, a number of in silico methods emerged to predict prokaryotic hosts based on viral sequences. One of the leading approaches is the application of the BLAST tool that searches for local similarities between viral and microbial genomes. However, this prediction method has three major limitations: (i) top-scoring sequences do not always point to the actual host; (ii) mosaic virus genomes may match to many, typically related, bacteria; and (iii) viral and host sequences may diverge beyond the point where their relationship can be detected by a BLAST alignment. Results We created an extension to BLAST, named Phirbo, that improves host prediction quality beyond what is obtainable from standard BLAST searches. The tool harnesses information concerning sequence similarity and bacteria relatedness to predict phage–host interactions. Phirbo was evaluated on three benchmark sets of known virus–host pairs, and it improved precision and recall by 11–40 percentage points over currently available, state-of-the-art, alignment-based, alignment-free, and machine-learning host prediction tools. Moreover, the discriminatory power of Phirbo for the recognition of virus–host relationships surpassed the results of other tools by at least 10 percentage points (area under the curve = 0.95), yielding a mean host prediction accuracy of 57% and 68% at the genus and family levels, respectively, and drops by 12 percentage points when using only a fraction of viral genome sequences (3 kb). Finally, we provide insights into a repertoire of protein and ncRNA genes that are shared between phages and hosts and may be prone to horizontal transfer during infection. Conclusions Our results suggest that Phirbo is a simple and effective tool for predicting phage–host relationships.

FishExp: a comprehensive database and analysis platform for gene expression and alternative splicing of fish species

The publicly archived RNA-seq data has grown exponentially, but its valuable information has not yet been fully discovered and utilized, especially for alternative splicing. This is true for fish species, which play important roles in ecology, research, and the food industry. To mitigate this, we present FishExp, a web-based data platform covering gene expression and alternative splicing in 26,081 RNA-seq experiments from 44 fishes. In addition to searching by gene identifiers and symbols, FishExp allows users to query the data using various functional terms and BLAST alignment. Notably, the user can customize experiments and tools to perform differential/specific expression and alternative splicing analysis, provided with functional enrichments. The results of retrieval and analysis can be visualized on the gene-, transcript- and splicing event-level webpage in a highly interactive and intuitive manner. The manually curated sample information, uniform data processing and visualization tools make it efficient for users to gain new insights from these large datasets. All data in FishExp can be downloaded for more in-depth analysis. FishExp is freely accessible at https://bioinfo.njau.edu.cn/fishExp.

Taxonomy-aware, sequence similarity ranking reliably predicts phage-host relationships

ABSTRACTMotivationSimilar regions in virus and host genomes provide strong evidence for phage-host interaction, and BLAST is one of the leading tools to predict hosts from phage sequences. However, BLAST-based host prediction has three limitations: (i) top-scoring prokaryotic sequences do not always point to the actual host, (ii) mosaic phage genomes may produce matches to many, typically related, bacteria, and (iii) phage and host sequences may diverge beyond the point where their relationship can be detected by a BLAST alignment.ResultsWe created an extension to BLAST, named Phirbo, that improves host prediction quality beyond what is obtainable from standard BLAST searches. The tool harnesses information concerning sequence similarity and bacteria relatedness to predict phage-host interactions. Phirbo was evaluated on two benchmark sets of known phage-host pairs, and it improved precision and recall by 25 percentage points, as well as the discriminatory power for the recognition of phage-host relationships by 10 percentage points (Area Under the Curve = 0.95). Phirbo also yielded a mean host prediction accuracy of 60% and 70% at the genus and family levels, respectively, representing a 5% improvement over BLAST. When using only a fraction of phage genome sequences (3 kb), the prediction accuracy of Phirbo was 5-11% higher than BLAST at all taxonomic levels.ConclusionOur results suggest that Phirbo is an effective, unsupervised tool for predicting phage-host relationships.AvailabilityPhirbo is available at https://github.com/aziele/phirbo.

Purification and Characterization of a Noble Thermostable Alpha-amylase from Anoxybacillus tengchongensis RA1-2-1 Isolated from Geothermal Spring of Nepal

A thermophilic amylolytic strain, Anoxybacillus tengchongensis RA1-2-1 was isolated from geothermal spring of Rasuwagadi district of Nepal. The BLAST alignment of the 16s rRNA sequence revealed 99.3% similarity with the type strain Anoxybacillus tengchongensis T-11. The morphological, physiological and biochemical properties were similar to the type strain. The enzyme from the strain was purified to 40-fold purification by DEAE-cellulose ion exchange chromatography. The Km value of the enzyme was 0.68±0.05 mg/ml. The optimum pH and temperature were 7.0 and 70 °C. SDS-PAGE analysis showed a single band at 69 kDa. The half-life of the enzyme at 70°C and 80°C were 85.01min and 51.96 min respectively. TLC analysis of the hydrolysis product showed that the enzyme is maltogenic amylase. The calcium independent enzyme was completely inhibited by Hg2+ but showed inhibitory effect in the range of 100 %-30 % in the presence of other salts at 1-10 mM concentrations.

PyFuncover: full proteome search for a specific function using BLAST and PFAM

Python Function uncover (PyFuncover) is a new bioinformatic tool able to search proteins with a specific function in a full proteome. The pipeline coded in python uses BLAST alignment and the sequences from a PFAM family as the search seed. We tested PyFuncover using the fatty acid-binding family (FABP) Lipocalin_7 from PFAM (version 32, 2019) against the Homo sapiens NCBI proteome. After applying the scoring function in all the BLAST results, the data were classified and submitted to a GO-TERM analysis using bioDBnet. Analyses showed that all families of FABPs were ranked within the top scores. Included within this category were also families able to bind to hydrophobic molecules similar to fatty acids such as the retinol acid transporter and the cellular retinoic acid-binding protein.

Proximate mechanisms involved in the formation of Secondary Osteon Morphotypes

SummaryProximate mechanisms involved in forming extracellular matrix (ECM) variations within and between bones are not yet clear. Deficiencies in the collective understanding of details required to illuminate the process that forms a highly ordered ECM are exposed when considering that there is still significant debate as to the importance of cellular control in the assembly of the ECM vs. the observation of collagen fibrillar “self-assembly” (i. e., occurring devoid of cells). We examined data and opinions with respect to possible mechanisms involved in the formation of distinctly different ECM patterns of secondary osteon morphotypes (SOMs). Important considerations include: (1) stretch within the osteoid during fibrillogenesis, (2) various mechanotransduction mechanisms, and (3) whether or not the formation of regional variations in osteonal ECMs requires osteo blast alignment and/or rotation and migration. We propose that primary cilia of osteoblasts and osteocytes have an important role in their perception of variant-related (vectorial) stimuli, which is deemed essential in the genesis of distinctive and mechanically relevant ECM patterns of SOMs.