Development of Sequence-Tagged Site Marker Set for Identification of J, JS, and St Sub-genomes of Thinopyrum intermedium in Wheat Background

Thinopyrum intermedium (2n = 6x = 42, JJJSJSStSt) is one of the important resources for the wheat improvement. So far, a few Th. intermedium (Thi)-specific molecular markers have been reported, but the number is far from enough to meet the need of identifying alien fragments in wheat-Th. intermedium hybrids. In this study, 5,877,409 contigs were assembled using the Th. intermedium genotyping-by-sequencing (GBS) data. We obtained 5,452 non-redundant contigs containing mapped Thi-GBS markers with less than 20% similarity to the wheat genome and developed 2,019 sequence-tagged site (STS) molecular markers. Among the markers designed, 745 Thi-specific markers with amplification products in Th. intermedium but not in eight wheat landraces were further selected. The distribution of these markers in different homologous groups of Th. intermedium varied from 47 (7/12/28 on 6J/6St/6JS) to 183 (54/62/67 on 7J/7St/7JS). Furthermore, the effectiveness of these Thi-specific markers was verified using wheat-Th. intermedium partial amphidiploids, addition lines, substitution lines, and translocation lines. Markers developed in this study provide a convenient, rapid, reliable, and economical method for identifying Th. intermedium chromosomes in wheat. In addition, this set of Thi-specific markers can also be used to estimate genetic and physical locations of Th. intermedium chromatin in the introgression lines, thus providing valuable information for follow-up studies such as alien gene mining.

De novo profiling of RNA viruses in Anopheles malaria vector mosquitoes from forest ecological zones in Senegal and Cambodia

BackgroundMosquitoes are colonized by a large but mostly uncharacterized natural virome of RNA viruses. Anopheles mosquitoes are efficient vectors of human malaria, and the composition and distribution of the natural RNA virome may influence the biology and immunity of Anopheles malaria vector populations.ResultsAnopheles vectors of human malaria were sampled in forest village sites in Senegal and Cambodia, including Anopheles funestus, Anopheles gambiae group sp., and Anopheles coustani in Senegal, and Anopheles hyrcanus group sp., Anopheles maculatus group sp., and Anopheles dirus in Cambodia. Small and long RNA sequences were depleted of mosquito host and de novo assembled to yield non-redundant contigs longer than 500 nucleotides. Analysis of the assemblies by sequence similarity to known virus families yielded 125 novel virus sequences, 39 from Senegal Anopheles and 86 from Cambodia. Important monophyletic virus clades in the Bunyavirales and Mononegavirales orders are found in these Anopheles from Africa and Asia. Small RNA size and abundance profiles were used to cluster non-host RNA assemblies that were unclassified by sequence similarity. 39 unclassified non-redundant contigs >500 nucleotides strongly matched a pattern of classic RNAi processing of viral replication intermediates, and 1566 unclassified contigs strongly matched a pattern consistent with piRNAs. Analysis of piRNA expression in Anopheles coluzzii after infection with O’nyong nyong virus (family Togaviridae) suggests that virus infection can specifically alter abundance of some piRNAs.ConclusionsRNA viruses ubiquitously colonize Anopheles vectors of human malaria worldwide. At least some members of the mosquito virome are monophyletic with other arthropod viruses. However, high levels of collinearity and similarity of Anopheles viruses at the peptide level is not necessarily matched by similarity at the nucleotide level, indicating that Anopheles from Africa and Asia are colonized by closely related but clearly diverged virome members. The interplay between small RNA pathways and the virome may represent an important part of the homeostatic mechanism maintaining virome members in a commensal or nonpathogenic state, and host-virome interactions could influence variation in malaria vector competence.

A Transcriptome Post-Scaffolding Method for Assembling High Quality Contigs

With the rapid development of high throughput sequencing technologies, new transcriptomes can be sequenced for little cost with high coverage. Sequence assembly approaches have been modified to meet the requirements for de novo transcriptomes, which have complications not found in traditional genome assemblies such as variation in coverage for each candidate mRNA and alternative splicing. As a consequence, de novo assembly strategies tend to generate a large number of redundant contigs due to sequence variations, which adversely affects downstream analysis and experiments. In this work we proposed TransPS, a transcriptome post-scaffolding method, to generate high quality, nonredundant de novo transcriptomes. TransPS shows promising results on the test transcriptome datasets, where redundancy is greatly reduced by more than 50% and, at the same time, coverage is improved considerably. The web server and source code are available.