Dataset Information

Variation in human chromosome 21 ribosomal RNA genes characterized by TAR cloning and long-read sequencing.

ABSTRACT: Despite the key role of the human ribosome in protein biosynthesis, little is known about the extent of sequence variation in ribosomal DNA (rDNA) or its pre-rRNA and rRNA products. We recovered ribosomal DNA segments from a single human chromosome 21 using transformation-associated recombination (TAR) cloning in yeast. Accurate long-read sequencing of 13 isolates covering ?0.82 Mb of the chromosome 21 rDNA complement revealed substantial variation among tandem repeat rDNA copies, several palindromic structures and potential errors in the previous reference sequence. These clones revealed 101 variant positions in the 45S transcription unit and 235 in the intergenic spacer sequence. Approximately 60% of the 45S variants were confirmed in independent whole-genome or RNA-seq data, with 47 of these further observed in mature 18S/28S rRNA sequences. TAR cloning and long-read sequencing enabled the accurate reconstruction of multiple rDNA units and a new, high-quality 44 838 bp rDNA reference sequence, which we have annotated with variants detected from chromosome 21 of a single individual. The large number of variants observed reveal heterogeneity in human rDNA, opening up the possibility of corresponding variations in ribosome dynamics.

SUBMITTER: Kim JH

PROVIDER: S-EPMC6061828 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature

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Variation in human chromosome 21 ribosomal RNA genes characterized by TAR cloning and long-read sequencing.

Kim Jung-Hyun JH Dilthey Alexander T AT Nagaraja Ramaiah R Lee Hee-Sheung HS Koren Sergey S Dudekula Dawood D Wood Iii William H WH Piao Yulan Y Ogurtsov Aleksey Y AY Utani Koichi K Noskov Vladimir N VN Shabalina Svetlana A SA Schlessinger David D Phillippy Adam M AM Larionov Vladimir V

Nucleic acids research 20180701 13

Despite the key role of the human ribosome in protein biosynthesis, little is known about the extent of sequence variation in ribosomal DNA (rDNA) or its pre-rRNA and rRNA products. We recovered ribosomal DNA segments from a single human chromosome 21 using transformation-associated recombination (TAR) cloning in yeast. Accurate long-read sequencing of 13 isolates covering ∼0.82 Mb of the chromosome 21 rDNA complement revealed substantial variation among tandem repeat rDNA copies, several palind ...[more]

PMID: 29788454

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Project description:MotivationLong arrays of near-identical tandem repeats are a common feature of centromeric and subtelomeric regions in complex genomes. These sequences present a source of repeat structure diversity that is commonly ignored by standard genomic tools. Unlike reads shorter than the underlying repeat structure that rely on indirect inference methods, e.g. assembly, long reads allow direct inference of satellite higher order repeat structure. To automate characterization of local centromeric tandem repeat sequence variation we have designed Alpha-CENTAURI (ALPHA satellite CENTromeric AUtomated Repeat Identification), that takes advantage of Pacific Bioscience long-reads from whole-genome sequencing datasets. By operating on reads prior to assembly, our approach provides a more comprehensive set of repeat-structure variants and is not impacted by rearrangements or sequence underrepresentation due to misassembly.ResultsWe demonstrate the utility of Alpha-CENTAURI in characterizing repeat structure for alpha satellite containing reads in the hydatidiform mole (CHM1, haploid-like) genome. The pipeline is designed to report local repeat organization summaries for each read, thereby monitoring rearrangements in repeat units, shifts in repeat orientation and sites of array transition into non-satellite DNA, typically defined by transposable element insertion. We validate the method by showing consistency with existing centromere high order repeat references. Alpha-CENTAURI can, in principle, run on any sequence data, offering a method to generate a sequence repeat resolution that could be readily performed using consensus sequences available for other satellite families in genomes without high-quality reference assemblies.Availability and implementationDocumentation and source code for Alpha-CENTAURI are freely available at http://github.com/volkansevim/alpha-CENTAURI CONTACT: ali.bashir@mssm.eduSupplementary informationSupplementary data are available at Bioinformatics online.

Dataset Information

Variation in human chromosome 21 ribosomal RNA genes characterized by TAR cloning and long-read sequencing.

Publications

Variation in human chromosome 21 ribosomal RNA genes characterized by TAR cloning and long-read sequencing.

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