Application of bioinformatics in assembling, annotating and drawing complete chloroplast genome of paphiopedilium delenatii (Guillaumin 1924) in Vietnam.pdf

NGUYEN TAT THANH UNIVERSITY NGUYEN TAT THANH True learning, true practice, true success, true future r FACULTY OF BIOTECHNOLOGY GRADUATION THESIS APPLICATION OF BIOINFORMATICS IN ASSEMBLING, ANNOTATING AND DRAWING COMPLETE CHLOROPLAST GENOME OF Paphiopedilum delenatii (Guillaumin 1924) IN VIETNAM Student’s name : Nguyen Thanh Diem Student ID : 1511540870 Major : Biotechnology Supervisors : MSc Vu Thi Huyen Trang Ho Chi Minh City, 2019 NGUYEN TAT THANH UNIVERSITY FACULTY OF BIOTECHNOLOGY SOCIALIST REPUBLIC OF VIETNAM Independence - Liberty - Happiness - oOo - ASSIGNED TASK OF GRADUATION THESIS Full name: Nguyen Thanh Diem Student ID: 1511540870 Major: Biotechnology Class: 15DSH1B Thesis’s title: Application of bioinformatics in assembling, annotating and drawing complete chloroplast genome of Paphiopedilunt delenatii (Guillaumin 1924) in Vietnam Objectives Application of bioinformatics in assembling, annotating and drawing complete chloroplast genome of the endemic species Paphiopedilum delenatii in Vietnam Contents - Extraction and quality assessment of DNA Raw data quality assessment Filtering out low quality sequences Genome assembly Genome annotation from contigs Construct genome map from annotated genome Execution time: from February/2018 to August/2019 Supervisor: MSc Vu Thi Huyen Trang Contents and requirements of this thesis were adopted by subject HCM city, ,2019 Subject Supervisor ACKNOWLEDGEMENTS To complete this graduation thesis, I would like thanks to my parents for always facilitating and encouraging me during my studies Sincere thanks to MSc Vu Thi Huyen Trang - Biotechnology Faculty - Nguyen Tat Thanh University for her whole heartedly guiding and creating all conditions for me during the process of thesis Sincerely thanks to: - Nguyen Tat Thanh University’s board of directors has created all conditions for me during my studies at the school - All teachers of Biotechnology Faculty - Nguyen Tat Thanh Unversity have taught me during the course at the school - My whole 15DSH1B class has helped me during my studies (Student’s signature) Nguyen Thanh Diem Faculty of Biotechnology Nguyen Tat Thanh University TABLE OF CONTENTS ACKNOWLEDGEMENTS i TABLE OF CONTENTS ii ABSTRACT V LIST OF FIGURES vi LIST OF TABLES vii LIST OF ACRONYMS viii INTRODUCTION ix CHAPTER LITERATURE REVIEW 1.1 Introduction of Paphiopedilum delenatii 1.1.1 Position of Paphiopedilum delenatii in the classification system 1.1.2 Morphological characteristics 1.1.3 Ecological characteristics 1.1.4 Individual status of Paphiopedilum in nature 1.2 Introduction of chloroplast genome 1.2.1 Introduction of chloroplast 1.2.2 Introduction of chloroplast genome 1.3 Introduction of bioinformatics and genome assembly .4 1.3.1 Application of Bioinformatics in genome assembly .4 1.3.2 Overview of genome annotation 1.3.3 Overview of genome map 1.4 Overview of softwares was used in this study 1.4.1 FastQC program 1.4.2 NOVOPlasty program ii 1.5 Related scientific research projects .9 1.5.1 Domestic projects 1.5.2 Foreign projects 10 CHAPTER CONTENTS AND METHODS 11 2.1 Place of administration 11 1.4.3 Chlorobox website 11 2.2 Contents 11 2.3 Methods 11 2.3.1 Materials 11 2.3.2 DNA extraction 11 2.3.3 Quality assessment of DNA 12 2.3.4 Quality assessment of raw data 12 2.3.5 Genome assembly 13 2.3.6 Genome annotation 14 2.3.7 Genome map construction 14 CHAPTER RESULTS 15 3.1 Results of DNA extraction 15 3.2 Raw data assessment 16 3.3 The results of genome assembly 18 3.4 The accuracy of genome structure 20 3.5 Genome annotation 21 3.6 The genome map construction 23 CHAPTER DISCUSSION 24 4.1 Raw data assessment 24 4.2 Phred quality score as input data for assembly 25 iii 4.3 The length of K-mer substring 25 4.4 The reference sequence (refseq) and seed sequence (seed) 26 4.5 Genome annotation 27 4.6 Genome map 29 CONCLUSIONS AND RECOMMENDATIONS 30 REFERENCES 31 IV ABSTRACT Chloroplasts and mitochondria are organelles that have their own genome The chloroplast genome provides information on the evolutionary relationship and identification of species, transgenic, cloning plants, etc Next Generation Sequencing makes it easier for chloroplast genome sequencing However, the assembly process of chloroplast genome is quite complicated due to the need of different complex bioinformatics tools, high configuration computer and time Here we configured the process of assembling the chloroplast genome of Paphiopedilum delenatii The assembled chloroplast genome was 160955 bp in length, including a large and a small single copy region (LSC, SSC) separated by a pair of inverted repeats (IR) Total genes were 130 genes, GC content was 35,6% Genome data was mapped and registered to GenBank as MK463585 The optimal parameters for genome assembling were recommended This study not only provided information for conservation of Vietnam endemic Paphiopedilum delenatii species but also supported the genome assemble researches which could be applied on other subjects V LIST OF FIGURES Figure 1.1 Morphology of Paphiopedilum delenatii Figure 1.2 Genome assembly steps Figure 1.3 De Bruijin graph Figure 1.4 Tire assembled chloroplast genome of p armeniacum Figure 1.5 Genome map of Citrus aurantiifolia Figure 3.1 The result of two p delenatii samples electrophoresis on 0.8 % 16 Figure 3.2 Quality assessment of raw data using FastQC 17 Figure 3.3 The BLAST result of two DNA sequence with reference genome of p armeniacum (KT388109.1) 20 Figure 3.4 The complete chloroplast genome map of Paphiopedilum delenatii 23 vi LIST OF TABLES Table 2.1 The length of reference sequences onNCBI 13 Table 2.2 The survey parameters were set for genome assembly process 13 Table 3.1 The results of OD and concentration of two p delenatii samples with Nanodrop and Quantus 15 Table 3.2 The reults of genome assembly 19 Table 3.3 List of annotated genes in chloroplast genome of p delenatii 22 Table 4.1 Evaluation of the correlation between quality score and reliability ratio 24 Table 4.2 Comparing p delenatii andP armeniacum genomes 28 vii LIST OF ACRONYMS bp : Base pair CR : Critically Endangered cp : Chlroplast NDH : NADH dehydrogenase - like complex NCBI : National Center for Biotechnology Information NGS : Next Generation Sequencing IUCN : International Union for Conservation of Nature IR : Invert repeat LSC : Larg single copy ssc : Small single copy viii ... title: Application of bioinformatics in assembling, annotating and drawing complete chloroplast genome of Paphiopedilunt delenatii (Guillaumin 1924) in Vietnam Objectives Application of bioinformatics. .. implemented the thesis ? ?Application of bioinformatics in assembling, annotating and drawing complete chloroplast genome of the endemic species Paphiopedilutn delenati (Guillaumin 1924) in Vietnam” IX... shedding light on the relationship between genomes of cp, mitochondria and nuclei in plants7 1.3 Introduction of bioinformatics and genome assembly 1.3.1 Application of Bioinformatics in genome