Iowa State University Capstones, Theses and Graduate Theses and Dissertations Dissertations 2019 Transcriptome analysis and identification of genes involved in moth sex pheromone biosynthetic pathways Xiaoyi Dou Iowa State University Follow this and additional works at: https://lib.dr.iastate.edu/etd Part of the Entomology Commons Recommended Citation Dou, Xiaoyi, "Transcriptome analysis and identification of genes involved in moth sex pheromone biosynthetic pathways" (2019). Graduate Theses and Dissertations. 17671. https://lib.dr.iastate.edu/etd/17671 This Dissertation is brought to you for free and open access by the Iowa State University Capstones, Theses and Dissertations at Iowa State University Digital Repository. It has been accepted for inclusion in Graduate Theses and Dissertations by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. Transcriptome analysis and identification of genes involved in moth sex pheromone biosynthetic pathways by Xiaoyi Dou A dissertation submitted to the graduate faculty in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Major: Entomology Program of Study Committee: Russell A. Jurenka, Major Professor Ryan C. Smith Joel R. Coats Amy L. Toth Hua Bai The student author, whose presentation of the scholarship herein was approved by the program of study committee, is solely responsible for the content of this dissertation. The Graduate College will ensure this dissertation is globally accessible and will not permit alterations after a degree is conferred Iowa State University Ames, Iowa 2019 Copyright © Xiaoyi Dou, 2019. All rights reserved. ii DEDICATION This study is dedicated to my family who have been my source of inspiration and give me strength. To my friends who give advice and encouragement to this study, thanks for your support. iii TABLE OF CONTENTS Page LIST OF FIGURES ....................................................................................................................... vi LIST OF TABLES ....................................................................................................................... viii ACKNOWLEDGMENTS ............................................................................................................. ix ABSTRACT .................................................................................................................................. xii CHAPTER 1. INTRODUCTION ................................................................................................... 1 Moths ........................................................................................................................................... 1 Sex Pheromone ............................................................................................................................ 3 Sex Pheromone Biosynthesis and Regulation ............................................................................. 5 Control Strategies using Sex Pheromones .................................................................................. 9 Dissertation Organization .......................................................................................................... 11 References ................................................................................................................................. 13 CHAPTER 2. TRANSCRIPTOME COMPARISON OF PHEROMONE GLAND- OVIPOSITOR AND TARSI IN THE CORN EARWORM MOTH, Helicoverpa zea ................ 20 Abstract ..................................................................................................................................... 20 Introduction ............................................................................................................................... 21 Materials and Methods .............................................................................................................. 23 Insects and tissue collections ................................................................................................. 23 RNA isolation, cDNA library construction and Illumina sequencing ................................... 23 Assembly of short reads and gene annotation ....................................................................... 24 Differential expression analysis ............................................................................................. 25 Identification of candidate genes involved in moth sex pheromone biosynthesis and chemosensation ...................................................................................................................... 25 Phylogenetic analysis ............................................................................................................ 26 Results ....................................................................................................................................... 26 Illumina sequencing, sequence assembly and gene annotation ............................................. 26 Differentially expressed genes ............................................................................................... 27 Candidate genes involved in sex pheromone biosynthesis .................................................... 28 Candidate genes involved in chemosensation and signal transduction ................................. 31 Discussion ................................................................................................................................. 33 Acknowledgements ................................................................................................................... 37 References ................................................................................................................................. 37 CHAPTER 3. IDENTIFICATION OF FATTY ACYL REDUCTASE IN PHEROMONE GLAND AND TARSI OF THE CORN EARWORM, Helicoverpa zea ..................................... 53 Abstract ..................................................................................................................................... 53 Introduction ............................................................................................................................... 54 Materials and Methods .............................................................................................................. 55 iv Insects .................................................................................................................................... 55 Chemicals .............................................................................................................................. 56 Cloning of the fatty acyl-CoA reductases .............................................................................. 56 RT-PCR ................................................................................................................................. 57 Functional Assay ................................................................................................................... 57 RNAi ...................................................................................................................................... 58 Quantitative real-time PCR ................................................................................................... 59 Phylogenetic construction ...................................................................................................... 60 Results ....................................................................................................................................... 60 Fatty acyl-CoA reductases cloning ........................................................................................ 60 RT-PCR indicates the three fatty acyl-CoA reductases are present in tarsi .......................... 61 Yeast expression and functional assay indicates fatty acyl-CoA reductases 1 produces fatty alcohols .................................................................................................................................. 62 RNAi indicates that FAR1 is involved in 16: Ald production in male tarsi .......................... 62 Discussion ................................................................................................................................. 63 Acknowledgments ..................................................................................................................... 65 References ................................................................................................................................. 65 CHAPTER 4. COMPARISON OF PINK BOLLWORM PHEROMONE GLAND TRANSCRIPTOME IN TWO POPULATIONS: LABORATORY AND FIELD ...................... 77 Abstract ..................................................................................................................................... 77 Introduction ............................................................................................................................... 78 Materials and Methods .............................................................................................................. 79 Insect collection and pheromone gland extraction and analysis ............................................ 79 RNA isolation, cDNA library construction and Illumina sequencing ................................... 80 De novo assembly of short reads and gene annotation .......................................................... 81 Expression abundance analysis ............................................................................................