Chang F.-M.,Taipei Medical University |
Ou T.-Y.,Taipei Medical University |
Cheng W.-N.,Taipei Medical University |
Chou M.-L.,Taipei Medical University |
And 6 more authors.
Fungal Genetics and Biology | Year: 2014
Candida albicans is considered to be an obligate diploid fungus. Here, we describe an approach to isolate aneuploids or haploids induced by the short-term (12-16. h) exposure of diploid reference strains SC5314 and CAI4 to the most commonly used antifungal drug, fluconazole, followed by repeated single-cell separation among small morphologically distinct colonies in the inhibition zone. The isolated strains had altered cell morphology and LOH events in the MTL and other marker alleles of the analyzed loci at 8 chromosomes of C. albicans with decreased DNA content. The present study employed next-generation sequencing (NGS) combined flow cytometry analysis of the DNA content to analyze the haploid, autodiploid, and aneuploid strains that arose from the fluconazole treatment instead of using the conventional single nucleotide polymorphism/comparative genome hybridization (SNP/CGH) method. A multiple-alignment tool was also developed based on sequenced data from NGS to establish haplotype mapping for each chromosome of the selected strains. These findings revealed that C. albicans experiences 'concerted chromosome loss' to form strains with homozygous alleles and that it even has a haploid status after short-term exposure to fluconazole. Additionally, we developed a new platform to analyze chromosome copy number using NGS. © 2014 Elsevier Inc.
Mohandas N.,University of Melbourne |
Pozio E.,Instituto Superiore Of Sanita |
La Rosa G.,Instituto Superiore Of Sanita |
Korhonen P.K.,University of Melbourne |
And 8 more authors.
International Journal for Parasitology | Year: 2014
In the present study we sequenced or re-sequenced, assembled and annotated 15 mitochondrial genomes representing the 12 currently recognised taxa of Trichinella using a deep sequencing-coupled approach. We then defined and compared the gene order in individual mitochondrial genomes (~14 to 17.7. kb), evaluated genetic differences among species/genotypes and re-assessed the relationships among these taxa using the mitochondrial nucleic acid or amino acid sequence data sets. In addition, a rich source of mitochondrial genetic markers was defined that could be used in future systematic, epidemiological and population genetic studies of Trichinella. The sequencing-bioinformatic approach employed herein should be applicable to a wide range of eukaryotic parasites. © 2014 Australian Society for Parasitology Inc.
Jayasundara D.,University of Melbourne |
Saeed I.,University of Melbourne |
Chang B.C.,Yourgene Bioscience |
Tang S.-L.,Academia Sinica, Taiwan |
Halgamuge S.K.,University of Melbourne
BMC Bioinformatics | Year: 2015
Background: Estimating the number of different species (richness) in a mixed microbial population has been a main focus in metagenomic research. Existing methods of species richness estimation ride on the assumption that the reads in each assembled contig correspond to only one of the microbial genomes in the population. This assumption and the underlying probabilistic formulations of existing methods are not useful for quasispecies populations where the strains are highly genetically related. The lack of knowledge on the number of different strains in a quasispecies population is observed to hinder the precision of existing Viral Quasispecies Spectrum Reconstruction (QSR) methods due to the uncontrolled reconstruction of a large number of in silico false positives. In this work, we formulated a novel probabilistic method for strain richness estimation specifically targeting viral quasispecies. By using this approach we improved our recently proposed spectrum reconstruction pipeline ViQuaS to achieve higher levels of precision in reconstructed quasispecies spectra without compromising the recall rates. We also discuss how one other existing popular QSR method named ShoRAH can be improved using this new approach. Results: On benchmark data sets, our estimation method provided accurate richness estimates (< 0.2 median estimation error) and improved the precision of ViQuaS by 2%-13% and F-score by 1%-9% without compromising the recall rates. We also demonstrate that our estimation method can be used to improve the precision and F-score of ShoRAH by 0%-7% and 0%-5% respectively. Conclusions: The proposed probabilistic estimation method can be used to estimate the richness of viral populations with a quasispecies behavior and to improve the accuracy of the quasispecies spectra reconstructed by the existing methods ViQuaS and ShoRAH in the presence of a moderate level of technical sequencing errors. © 2015 Jayasundara et al.
Jayasundara D.,Optimisation and Pattern Recognition Research Group |
Saeed I.,Optimisation and Pattern Recognition Research Group |
Maheswararajah S.,Portland House Research and Advisors Ltd. |
Chang B.C.,Yourgene Bioscience |
And 2 more authors.
Bioinformatics | Year: 2015
Motivation: The combined effect of a high replication rate and the low fidelity of the viral polymerase in most RNA viruses and some DNA viruses results in the formation of a viral quasispecies. Uncovering information about quasispecies populations significantly benefits the study of disease progression, antiviral drug design, vaccine design and viral pathogenesis. We present a new analysis pipeline called ViQuaS for viral quasispecies spectrum reconstruction using short next-generation sequencing reads. ViQuaS is based on a novel reference-assisted de novo assembly algorithm for constructing local haplotypes. A significantly extended version of an existing global strain reconstruction algorithm is also used. Results: Benchmarking results showed that ViQuaS outperformed three other previously published methods named ShoRAH, QuRe and PredictHaplo, with improvements of at least 3.1-53.9% in recall, 0-12.1% in precision and 0-38.2% in F-score in terms of strain sequence assembly and improvements of at least 0.006-0.143 in KL-divergence and 0.001-0.035 in root mean-squared error in terms of strain frequency estimation, over the next-best algorithm under various simulation settings. We also applied ViQuaS on a real read set derived from an in vitro human immunodeficiency virus (HIV)-1 population, two independent datasets of foot-and-mouth-disease virus derived from the same biological sample and a real HIV-1 dataset and demonstrated better results than other methods available. Availability and implementation: http://sourceforge.net/projects/viquas/. © The Author 2014. Published by Oxford University Press. All rights reserved.
Yourgene Bioscience | Date: 2015-03-03
A method for detecting a chromosomal aneuploidy relating to a target nucleic acid region includes the following steps. A reference database is obtained. At least one normalizing factor is determined based on the reference database. A cutoff value is determined based on the reference database. A biological sample under test is sequenced by the sequencing platform to obtain a number of target reads of the biological sample under test. The target reads of the biological sample under test originate from the target nucleic acid region. The number of the target reads of the biological sample under test is normalized by the normalizing factor and then is compared with the cutoff value. Whether the chromosomal aneuploidy relating to the target nucleic acid region is present in the fetus is determined based on the comparison
Yang S.-H.,Academia Sinica, Taiwan |
Yang S.-H.,National Chung Hsing University |
Lee S.T.M.,University of Auckland |
Huang C.-R.,Academia Sinica, Taiwan |
And 6 more authors.
Limnology and Oceanography | Year: 2016
Microbial endoliths, which inhabit interior pores of rocks, skeletons and coral, are ubiquitous in terrestrial and marine environments. In this study, various colored layers stratified the endolithic environment within the skeleton of Isopora palifera; however, there was a distinct green-pigmented layer in the skeleton (beneath the living coral tissue). To characterize diversity of endolithic microorganisms, 16S ribosomal RNA gene amplicon pyrosequencing was used to investigate bacterial communities in the green layer of eight I. palifera coral colonies retrieved from two locations on Green Island, Taiwan. The dominant bacterial group in the green layer belonged to the bacterial phylum Chlorobi, green sulfur bacteria capable of anoxygenic photosynthesis and nitrogen fixation. Specifically, bacteria of the genus Prosthecochloris were dominant in this green layer. To our knowledge, this is the first study to provide a detailed profile of endolithic bacteria in coral and to determine the prevalence of Prosthecochloris in the green layer. Based on our findings, we infer that these bacteria may have an important functional role in the coral holobiont in the nutrient-limited coral reef ecosystem. © 2016 Association for the Sciences of Limnology and Oceanography.
Yourgene Bioscience | Date: 2016-01-27
A method for detecting a chromosomal aneuploidy relating to a target nucleic acid region includes the following steps. A reference database is obtained. At least one normalizing factor is determined based on the reference database. A cutoff value is determined based on the reference database. A biological sample under test is sequenced by the sequencing platform to obtain a number of target reads of the biological sample under test. The target reads of the biological sample under test originate from the target nucleic acid region. The number of the target reads of the biological sample under test is normalized by the normalizing factor and then is compared with the cutoff value. Whether the chromosomal aneuploidy relating to the target nucleic acid region is present in the fetus is determined based on the comparison.
PubMed | National Cheng Kung University and Yourgene Bioscience
Type: Journal Article | Journal: Hepatology international | Year: 2016
Hepatitis B virus (HBV) quasispecies are crucial in the pathogenesis of chronic liver disease. Next-generation sequencing (NGS) is powerful for identifying viral quasispecies. To improve mapping quality and single nucleotide variant (SNV) calling accuracy in the NGS analysis of HBV, we compared different mapping references, including the sample-specific reference sequence, same genotype sequences and different genotype sequences, according to the sample.Real Illumina HBV datasets from 86 patients, and simulated datasets from 158 HBV strains in the GenBank database, were used to assess mapping quality. SNV calling accuracy was evaluated using different mapping references to align Real Illumina datasets from a single HBV clone.Using the sample-specific reference sequence as a mapping reference produced the largest number of mappable reads and coverages. With a different genotype mapping reference, the consensus sequence derived from the Real Illumina datasets of the single HBV clone showed 21 false SNV callings in polymerase and surface genes, the regions most divergent between the mapping reference and this HBV clone. A ~6% coverage of most of these false SNVs was yielded even with a same genotype mapping reference, but none with the sample-specific reference sequence.Using sample-specific reference sequences as a mapping reference in NGS analysis optimized mapping quality and the SNV calling accuracy for HBV quasispecies.
PubMed | Yourgene Bioscience and Academia Sinica, Taiwan
Type: Journal Article | Journal: G3 (Bethesda, Md.) | Year: 2014
Coconut palm (Cocos nucifera) is a symbol of the tropics and a source of numerous edible and nonedible products of economic value. Despite its nutritional and industrial significance, coconut remains under-represented in public repositories for genomic and transcriptomic data. We report de novo transcript assembly from RNA-seq data and analysis of gene expression in seed tissues (embryo and endosperm) and leaves of a dwarf coconut variety. Assembly of 10 GB sequencing data for each tissue resulted in 58,211 total unigenes in embryo, 61,152 in endosperm, and 33,446 in leaf. Within each unigene pool, 24,857 could be annotated in embryo, 29,731 could be annotated in endosperm, and 26,064 could be annotated in leaf. A KEGG analysis identified 138, 138, and 139 pathways, respectively, in transcriptomes of embryo, endosperm, and leaf tissues. Given the extraordinarily large size of coconut seeds and the importance of small RNA-mediated epigenetic regulation during seed development in model plants, we used homology searches to identify putative homologs of factors required for RNA-directed DNA methylation in coconut. The findings suggest that RNA-directed DNA methylation is important during coconut seed development, particularly in maturing endosperm. This dataset will expand the genomics resources available for coconut and provide a foundation for more detailed analyses that may assist molecular breeding strategies aimed at improving this major tropical crop.
PubMed | Ming Chuan University, National Pingtung University of Science and Technology, University of Melbourne and Yourgene Bioscience
Type: | Journal: PeerJ | Year: 2016
The Phalaenopsis orchid is an important potted flower of high economic value around the world. We report the 3.1 Gb draft genome assembly of an important winter flowering Phalaenopsis KHM190 cultivar. We generated 89.5 Gb RNA-seq and 113 million sRNA-seq reads to use these data to identify 41,153 protein-coding genes and 188 miRNA families. We also generated a draft genome for Phalaenopsis pulcherrima B8802, a summer flowering species, via resequencing. Comparison of genome data between the two Phalaenopsis cultivars allowed the identification of 691,532 single-nucleotide polymorphisms. In this study, we reveal that the key role of PhAGL6b in the regulation of labellum organ development involves alternative splicing in the big lip mutant. Petal or sepal overexpressing PhAGL6b leads to the conversion into a lip-like structure. We also discovered that the gibberellin pathway that regulates the expression of flowering time genes during the reproductive phase change is induced by cool temperature. Our work thus depicted a valuable resource for the flowering control, flower architecture development, and breeding of the Phalaenopsis orchids.