Evaluation of genetic diversity of some dendrobium orchids in southern vietnam

NGUYEN TAT THANH UNIVERSITY TRUE LEARNING - TRUE PRACTICE - TRUE SUCCESS - TRUE FUTURE FACULTY OF BIOTECHNOLOGY GRADUATION THESIS EVALUATION OF GENETIC DIVERSITY OF SOME DENDROBIUM ORCHIDS IN SOUTHERN VIETNAM Student’s name : Le Thi Ngoc Diep Student ID : 1711547413 Supervisor : MSc Vu Thi Huyen Trang Ho Chi Minh City, 2020 NGUYEN TAT THANH UNIVERSITY TRUE LEARNING - TRUE PRACTICE - TRUE SUCCESS - TRUE FUTURE FACULTY OF BIOTECHNOLOGY GRADUATION THESIS EVALUATION OF GENETIC DIVERSITY OF SOME DENDROBIUM ORCHIDS IN SOUTHERN VIETNAM Student’s name : Le Thi Ngoc Diep Student ID : 1711547413 Major : Biotechnology Supervisors : MSc Vu Thi Huyen Trang Ho Chi Minh City, 2020 NGUYEN TAT THANH UNIVERSITY SOCIALIST REPUBLIC OF VIETNAM FACULTY OF BIOTECHNOLOGY Independence – Liberty - Happiness -oOo - ASSIGNED TASK OF GRADUATION THESIS Full name: Le Thi Ngoc Diep Student ID: 1711547413 Major: Biotechnology Class: 17DSH1A Thesis’s title: Evaluation of genetic diversity of some Dendrobium orchids in southern Vietnam Objectives - Constructing ITS and matK sequence data set of some Dendrobium varieties - Evaluation of genetic diversity of Dendrobium varieties in the Southern Vietnam based on two regions of sequence ITS and matK Contents - Sampling, extracting total DNA and amplifying two sequence regions matK and ITS - Sequence editing and alignment - Species identification and analysis of genetic diversity Execution time: from 06/2020 to 09/2020 Supervisor: MSc Vu Thi Huyen Trang Contents and requirements of this thesis were adopted by subject HCM city, September 20, 2020 Faculty/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 wholeheartedly 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 aught me during the course at the school − My whole 17DSH1A class has helped me during my studies Le Thi Ngoc Diep Faculty of Biotechnology Nguyen Tat Thanh University i TABLE OF CONTENTS ACKNOWLEDGEMENTS i TABLE OF CONTENTS ii ABSTRACT iv LIST OF FIGURES .v LIST OF TABLES vi LIST OF ACRONYMS vii INTRODUCTION viii CHAPTER LITERATURE REVIEW 1.1 Introduction to the family Orchidaceae 1.2 Introduction of Dendrobium genus 1.2.1 Scientific classification 1.2.2 Distribution and habitat 1.2.3 Morphology of Dendrobium 1.2.4 Habitat 1.2.5 The role and value of Dendrobium .5 1.3 Introduction to genetic diversity .6 1.3.1 Concept 1.3.2 Role of genetic diversity analysis 1.3.3 Methods of genetic diversity analysis 1.4 Research situation 1.4.1 International studies 1.4.2 Viet Nam researchs .11 CHAPTER CONTENTS AND METHODS 12 2.1 Place of administration 12 ii 2.2 Research content 12 2.3 Research methodology 12 2.3.1 Research materials 12 2.3.2 Total DNA extraction, amplification of ITS and matK sequence regions 12 CHAPTER RESULTS AND DISCUSSION 14 3.1 Sample collection 14 3.2 Amplification and sequence adjustment .15 3.3 Genetic diversity of Dendrobium species .17 3.4 Discussion .22 CONCLUSIONS AND RECOMMENDATIONS 23 Conclusions 23 Recommendations 23 REFERENCES 24 APPENDICES 27 iii ABSTRACT Thesis title: "Evaluation of genetic diversity of some Dendrobium orchids in southern Vietnam" Dendrobium is the genus with large number of species and diversity in morphology and colors In Vietnam, there are about 107 Dendrobium species, which widely distributed in mountainous areas from North to South and even in some coastal areas Due to their diversity, morphological recognition of Dendrobium taxa is still in challenges which leads to misidentification among species Since then, diversity assessment based on molecular genetic feature helps in exact identification, serving for the conservation of species, management of seed breeding, and hybridization for creating new valuable variations In this study, we conducted genetic diversity evaluation of 30 Dendrobium samples in South Vietnam The results identified the exact scientific names of 16 species The ITS region gave high potential in identifying this orchid group This study also contributes to the identification, selection and breeding of new varieties of Dendrobium species in Vietnam iv LIST OF FIGURES Figure 1.1 Dendrobium suzukii species Figure 3.1 Amplification results of ITS region .15 Figure 3.2 Amplification results of matK region 16 Figure 3.3 Phylogenetic tree based on ITS sequence 18 Figure 3.4 Phylogenetic tree based on matK sequence 19 Figure 3.5 Morphology of stems and leaves of D hancockii (A) and D saclaccence (B) 20 Figure 3.6 Similarity search based on ITS of 24DT sample matched to D hancockii 21 Figure 3.7 Similarity search based on matK of 24DT sample matched to D hancockii .21 v LIST OF TABLES Table 2.1 Primer sequences for amplification reactions of ITS, matK regions 13 Table 2.2 Thermal cycle of PCR reaction of ITS 13 Table 2.3 Thermal cycle of PCR reaction of matK .13 Table 3.1 30 samples of Dendrobium in the study 14 Table 3.2 The results of DNA extraction and sequencing 17 Table 3.3 Diversity of Dendrobium species from the study 21 vi LIST OF ACRONYMS AFLP: Amplified Fragment LengthPolymophism ITS: Internal Transcribed Spacer SSCP: Single Strand Conformation Polymophism SSR: Simple Sequence Repeat PCR: Polymerase Chain Reaction RAPD: Random Amplified Polymorphic DNA RFLP: Restriction Fragment Length Polymophism vii ... title: Evaluation of genetic diversity of some Dendrobium orchids in southern Vietnam Objectives - Constructing ITS and matK sequence data set of some Dendrobium varieties - Evaluation of genetic diversity. .. title: "Evaluation of genetic diversity of some Dendrobium orchids in southern Vietnam" Dendrobium is the genus with large number of species and diversity in morphology and colors In Vietnam, ... manage genetic diversity and conserve rare valuable orchids Since then, we performed the project: "Evaluation of genetic diversity of some Dendrobium orchids in southern Vietnam" to determine the genetic