Dataset Information

Building two indica rice reference genomes with PacBio long-read and Illumina paired-end sequencing data.

ABSTRACT: Over the past 30 years, we have performed many fundamental studies on two Oryza sativa subsp. indica varieties, Zhenshan 97 (ZS97) and Minghui 63 (MH63). To improve the resolution of many of these investigations, we generated two reference-quality reference genome assemblies using the most advanced sequencing technologies. Using PacBio SMRT technology, we produced over 108 (ZS97) and 174 (MH63) Gb of raw sequence data from 166 (ZS97) and 209 (MH63) pools of BAC clones, and generated ~97 (ZS97) and ~74 (MH63) Gb of paired-end whole-genome shotgun (WGS) sequence data with Illumina sequencing technology. With these data, we successfully assembled two platinum standard reference genomes that have been publicly released. Here we provide the full sets of raw data used to generate these two reference genome assemblies. These data sets can be used to test new programs for better genome assembly and annotation, aid in the discovery of new insights into genome structure, function, and evolution, and help to provide essential support to biological research in general.

SUBMITTER: Zhang J

PROVIDER: S-EPMC5020871 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature

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Building two indica rice reference genomes with PacBio long-read and Illumina paired-end sequencing data.

Zhang Jianwei J Chen Ling-Ling LL Sun Shuai S Kudrna Dave D Copetti Dario D Li Weiming W Mu Ting T Jiao Wen-Biao WB Xing Feng F Lee Seunghee S Talag Jayson J Song Jia-Ming JM Du Bogu B Xie Weibo W Luo Meizhong M Maldonado Carlos Ernesto CE Goicoechea Jose Luis JL Xiong Lizhong L Wu Changyin C Xing Yongzhong Y Zhou Dao-Xiu DX Yu Sibin S Zhao Yu Y Wang Gongwei G Yu Yeisoo Y Luo Yijie Y Hurtado Beatriz Elena Padilla BE Danowitz Ann A Wing Rod A RA Zhang Qifa Q

Scientific data 20160913

Over the past 30 years, we have performed many fundamental studies on two Oryza sativa subsp. indica varieties, Zhenshan 97 (ZS97) and Minghui 63 (MH63). To improve the resolution of many of these investigations, we generated two reference-quality reference genome assemblies using the most advanced sequencing technologies. Using PacBio SMRT technology, we produced over 108 (ZS97) and 174 (MH63) Gb of raw sequence data from 166 (ZS97) and 209 (MH63) pools of BAC clones, and generated ~97 (ZS97) a ...[more]

PMID: 27622467

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Project description:BACKGROUND:Illumina sequencing of a marker gene is popular in metagenomic studies. However, Illumina paired-end (PE) reads sometimes cannot be merged into single reads for subsequent analysis. When mergeable PE reads are limited, one can simply use only first reads for taxonomy annotation, but that wastes information in the second reads. Presumably, including second reads should improve taxonomy annotation. However, a rigorous investigation of how best to do this and how much can be gained has not been reported. RESULTS:We evaluated two methods of joining as opposed to merging PE reads into single reads for taxonomy annotation using simulated data with sequencing errors. Our rigorous evaluation involved several top classifiers (RDP classifier, SINTAX, and two alignment-based methods) and realistic benchmark datasets. For most classifiers, read joining ameliorated the impact of sequencing errors and improved the accuracy of taxonomy predictions. For alignment-based top-hit classifiers, rearranging the reference sequences is recommended to avoid improper alignments of joined reads. For word-counting classifiers, joined reads could be compared to the original reference for classification. We also applied read joining to our own real MiSeq PE data of nasal microbiota of asthmatic children. Before joining, trimming low quality bases was necessary for optimizing taxonomy annotation and sequence clustering. We then showed that read joining increased the amount of effective data for taxonomy annotation. Using these joined trimmed reads, we were able to identify two promising bacterial genera that might be associated with asthma exacerbation. CONCLUSIONS:When mergeable PE reads are limited, joining them into single reads for taxonomy annotation is always recommended. Reference sequences may need to be rearranged accordingly depending on the classifier. Read joining also relaxes the constraint on primer selection, and thus may unleash the full capacity of Illumina PE data for taxonomy annotation. Our work provides guidance for fully utilizing PE data of a marker gene when mergeable reads are limited.

Project description:BackgroundLarge insert paired-end sequencing technologies are important tools for assembling genomes, delineating associated breakpoints and detecting structural rearrangements. To facilitate the comprehensive detection of inter- and intra-chromosomal structural rearrangements or variants (SVs) and complex genome assembly with long repeats and segmental duplications, we developed a new method based on single-molecule real-time synthesis sequencing technology for generating long paired-end sequences of large insert DNA libraries.ResultsA Fosmid vector, pHZAUFOS3, was developed with the following new features: (1) two 18-bp non-palindromic I-SceI sites flank the cloning site, and another two sites are present in the skeleton of the vector, allowing long DNA inserts (and the long paired-ends in this paper) to be recovered as single fragments and the vector (~?8 kb) to be fragmented into 2-3 kb fragments by I-SceI digestion and therefore was effectively removed from the long paired-ends (5-10 kb); (2) the chloramphenicol (Cm) resistance gene and replicon (oriV), necessary for colony growth, are located near the two sides of the cloning site, helping to increase the proportion of the paired-end fragments to single-end fragments in the paired-end libraries. Paired-end libraries were constructed by ligating the size-selected, mechanically sheared pooled Fosmid DNA fragments to the Ampicillin (Amp) resistance gene fragment and screening the colonies with Cm and Amp. We tested this method on yeast and Setaria italica Yugu1. Fosmid-size paired-ends with an average length longer than 2 kb for each end were generated. The N50 scaffold lengths of the de novo assemblies of the yeast and S. italica Yugu1 genomes were significantly improved. Five large and five small structural rearrangements or assembly errors spanning tens of bp to tens of kb were identified in S. italica Yugu1 including deletions, inversions, duplications and translocations.ConclusionsWe developed a new method for long paired-end sequencing of large insert libraries, which can efficiently improve the quality of de novo genome assembly and identify large and small structural rearrangements or assembly errors.

Project description:BackgroundMiscanthus sinensis Andersson is a perennial grass that exhibits remarkable lignocellulose characteristics suitable for sustainable bioenergy production. However, knowledge of the genetic resources of this species is relatively limited, which considerably hampers further work on its biology and genetic improvement.ResultsIn this study, through analyzing the transcriptome of mixed samples of leaves and stems using the latest PacBio Iso-Seq sequencing technology combined with Illumina HiSeq, we report the first full-length transcriptome dataset of M. sinensis with a total of 58.21 Gb clean data. An average of 15.75 Gb clean reads of each sample were obtained from the PacBio Iso-Seq system, which doubled the data size (6.68 Gb) obtained from the Illumina HiSeq platform. The integrated analyses of PacBio- and Illumina-based transcriptomic data uncovered 408,801 non-redundant transcripts with an average length of 1,685 bp. Of those, 189,406 transcripts were commonly identified by both methods, 169,149 transcripts with an average length of 619 bp were uniquely identified by Illumina HiSeq, and 51,246 transcripts with an average length of 2,535 bp were uniquely identified by PacBio Iso-Seq. Approximately 96 % of the final combined transcripts were mapped back to the Miscanthus genome, reflecting the high quality and coverage of our sequencing results. When comparing our data with genomes of four species of Andropogoneae, M. sinensis showed the closest relationship with sugarcane with up to 93 % mapping ratios, followed by sorghum with up to 80 % mapping ratios, indicating a high conservation of orthologs in these three genomes. Furthermore, 306,228 transcripts were successfully annotated against public databases including cell wall related genes and transcript factor families, thus providing many new insights into gene functions. The PacBio Iso-Seq data also helped identify 3,898 alternative splicing events and 2,963 annotated AS isoforms within 10 function categories.ConclusionsTaken together, the present study provides a rich data set of full-length transcripts that greatly enriches our understanding of M. sinensis transcriptomic resources, thus facilitating further genetic improvement and molecular studies of the Miscanthus species.

Dataset Information

Building two indica rice reference genomes with PacBio long-read and Illumina paired-end sequencing data.

Publications

Building two indica rice reference genomes with PacBio long-read and Illumina paired-end sequencing data.

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