Generation of reference genomes for Trypanosoma cruzi using PacBio sequencing

使用 PacBio 测序生成克氏锥虫参考基因组

• 批准号: 9251753
• 负责人: Rick L Tarleton
• 金额: $ 7.5万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9251753
• 关键词: Address; Americas; Biological Phenomena; Blood; Cells; Chagas Disease; Chromosomes; Communities; Complex; Consensus; Data; Dideoxy Chain Termination DNA Sequencing; Diploidy; Epidemiology; Etiology; Evolution; Future; Gene Duplication; Gene Family; Generations; Genes; Genetic; Genome; Genome Mappings; Genomic DNA; Genomics; Goals; Grant; Heterozygote; Hybrids; Immune Evasion; Institutes; Ions; Karyotype; Laboratories; Length; Libraries; Maintenance; Manuals; Maps; Membrane Proteins; Modification; Molecular Weight; Mosaicism; Nature; Outcome; PPBP gene; Parasites; Pharmacotherapy; Prevention; Protons; Structure; Technology; Testing; Trypanosoma cruzi; Validation; base; duplicate genes; genome sequencing; genome-wide; human disease; improved; interest; next generation sequencing; programs; public health relevance; reference genome; single molecule; virtual; whole genome

 DESCRIPTION (provided by applicant): The most persistent and significant hindrance to genomic studies of Trypanosoma cruzi is the lack of an acceptable reference genome. There are a number of factors that have impeded progress in generating a high quality, fully assembled T. cruzi reference sequence, including high repeat content (50%) in the T. cruzi genome, extreme genome-wide heterozygosity in the reference strain (CL-Brener) that was chosen for sequencing and reference genome assembly, and genetic mosaicism in the reference strain. Thus, the current reference genome for T. cruzi has hundreds of gaps and regions of assembly collapse, has never been fully assembled and is, in fact, presented as two genomes due to the heterozygosity of the reference strain -each chromosome is presented as two versions differing substantially in terms of gene content, sequence identity where the two chromosome sequences align, and even lengths of the two chromosomes in a pair. These issues have greatly muted the efficacy and power of previous and ongoing whole genome sequencing studies in this causative agent of human disease, and have made virtually any endeavor involving the need for scrutinizing the reference genome difficult and frequently unproductive. We propose to tackle the issues with the current T. cruzi reference genome using three modifications of the approach originally used to generate it. First, we propose to use single-molecule, long-read (up to 10 kb) sequencing to close gaps and expand regions of assembly collapse that occurred when primarily Sanger reads (700 bp) were used to assemble the original CL-Brener reference genome. Second, we will select T. cruzi isolates for sequencing that are homozygous and do not have mosaic genomes to simplify the assemblies. Third, because all T. cruzi lines are derived from two ancestral, divergent, homozygous, non-mosaic lineages, we will generate reference sequences to each "parental" lineage in order to cover the scope of genetic content in all T. cruzi isolates.