Publications

Detailed Information

Development of Optimized Bioinformatic Analysis Systems for Next Generation Sequencing Data from Metagenome : 차세대 염기서열 분석 장비로 생성한 메타지놈 데이터 분석을 위한 최적의 생물정보학 시스템 개발

Cited 0 time in Web of Science Cited 0 time in Scopus
Authors

전윤성

Advisor
천종식
Major
자연과학대학 협동과정 생물정보학전공
Issue Date
2014-02
Publisher
서울대학교 대학원
Keywords
bioinformaticsnext generation sequencingmetagenomeampliconshotgun metagenomeanalysis pipelineEzEditorphylogenetics
Description
학위논문 (박사)-- 서울대학교 대학원 : 협동과정 생물정보학전공, 2014. 2. 천종식.
Abstract
Metagenome is total DNA directly extracted from environment, and the purpose of metagenomics is to reveal the function of the metagenome as well as the taxonomic structure in the metagenome. There are two analysis approaches for metagenomics, namely amplicon based approach and random shotgun based approach. Both approaches require large scale sequencing reads which could not be satisfied through Sanger sequencing. However, high throughput sequencing of reads at relatively low cost by Next Generation Sequencing (NGS) technologies meets the requirement of metagenomics. In addition, the advent of NGS technologies gave rise to the development of bioinformatic algorithms necessary for processing this large and complex sequencing data. Consequently, the large amount of sequencing data obtained from NGS and corresponding proper bioinformatic algorithms facilitated the metagenomics to become essential tool for microbiology. However, limitations incurred by NGS sequencing errors, short read length, and lack of analysis system still hinder accurate metagenome analysis. Therefore, evaluation of currently used NGS error handling algorithms and development of systematic pipeline with more efficient algorithms are required to improve the accuracy of analysis.
In this study, bioinformatic pipelines were constructed for both metagenome analysis approaches. The pipelines were dedicated to improve the accuracy of the final end result by minimizing the effect of errors and short read length. For the amplicon based metagenomics, two different analysis pipelines were developed for both 454 pyrosequencing and Illumina MiSeq. During the construction of 454 pyrosequencing pipeline, new error handling algorithm was developed to treat homo-polymer and PCR errors. Upon completion of the pipeline construction, household microbial community was analyzed using 454 pyrosequencing data as a case study. As for Illumina MiSeq data, the most appropriate sequencing conditions and sequencing target region were settled. Paired end merging programs were evaluated and correlation of the sequencing errors and quality was studied to correct the errors within 3 overlap regions. Novel iterative consensus clustering method was developed to correct the errors occurring ubiquitously in a single read.
For shotgun metagenomics approach, bioinformatic analysis system for Illumina MiSeq paired end data was constructed. Unlike the targeted amplicon sequencing reads, most of the shotgun sequencing reads are not merged
thus short reads are used for both functional and taxonomical profiling. However, a short read has less information than longer contigs, so the use of short reads is likely to cause biased characterization of the metagenome. Therefore, the development of analysis system did focus on creating longer contigs by means of mapping and de novo assembly. For raw read mapping, a dynamic mapping genome set construction method was developed. A list of mapping genomes was selected from the taxonomic profile inferred from the ribosomal RNA profiles. The genome sequence of the selected genomes were downloaded from Ezbiocloud. By mapping raw reads to the genome sequences, the longer contigs can be obtained in case of the relatively simple metagenome such as fecal matter. However in case of the complex metagenomes such as soil sample, both mapping and de novo assembly did not perform properly due to a lack of sequencing coverage and numerousity of uncultured microorganisms in the metagenome. In addition to the pipeline construction, visualization tools were also developed to display resultant taxonomic and functional profile at the same time.
Newly developed JAVA-based standalone sequence alignment editing application was named as EzEditor. As both, conserved functional coding sequences and 16S rRNA gene have been used copiously in bacterial molecular phylogenetics, the codon-based sequence alignment editing functions are required for the coding genes. EzEditor provides simultaneous DNA and protein sequence alignment editing interface which enables us with the robust sequence alignment for both protein and rRNA sequences. EzEditor can be applied to various molecular sequence involved analysis not only as a basic sequence editor but also for phylogenetic application.
Language
Korean
URI
https://hdl.handle.net/10371/125376
Files in This Item:
Appears in Collections:

Altmetrics

Item View & Download Count

  • mendeley

Items in S-Space are protected by copyright, with all rights reserved, unless otherwise indicated.

Share