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Bacterial Diversity in the Rhizosphere of AVP1 Transgenic Cotton (Gossypium Hirsutum L.) and Wheat (Triticum Aestivum L.)

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Bacterial Diversity in the Rhizosphere of AVP1 Transgenic Cotton (Gossypium Hirsutum L.) and Wheat (Triticum Aestivum L.) Bacterial Diversity in the Rhizosphere of AVP1 Transgenic Cotton (Gossypium hirsutum L.) and Wheat (Triticum aestivum L.) Muhammad Arshad 2016 Department of Biotechnology Pakistan Institute of Engineering & Applied Sciences Nilore-45650 Islamabad, Pakistan Reviewers and Examiners Foreign Reviewers 1. Dr. Dittmar Hahn Department of Biology, Texas State University, 601 University Drive San Marcos, Fax +1 (512) 245 8713 Telephone: +1 (512) 245 3372 E-mail Address: [email protected] 2. Dr. Philippe Normad Microbial Ecology Laboratory, UMR CNRS 5557, F-69622 Villeurbanne Cedex Telephone: 33 (0)4-7243-1377 E-mail Address: [email protected] University of Arkansas, 3. Dr. Katharina Pawlowski Stockholm University Mailing Address: SE-106 91 Stockholm, Sweden Telephone (With Country Code): +46 8 16 37 72 E-mail Address: [email protected] Thesis Examiners 1. Dr. Asghari Bano, Department of Biosciences university of Wah cant Telephone # 03129654341 E-mail Address: [email protected] 2. Dr. Muhammad Arshad Department of Botany, PMAS AAU, Murree Road, Rawalpindi Telephone: 051-9062207 E-mail Address: [email protected] 3. Dr. Amer Jamil, Molecular Biochemistry Lab, Dept. of Chemistry and Biochemistry, University of Agriculture Faisalabad Telephone: 41-9201104 E-mail Address: [email protected] Head of the Department (Name): Prof. Dr. Shahid Mansoor, S.I. Signature with date: _____________________ Thesis Submission Approval This is to certify that the work contained in this thesis entitled Bacterial Diversity in the Rhizosphere of AVP1 Transgenic Cotton (Gossypium hirsutum L.) and Wheat (Triticum aestivum L.), was carried out by Muhammad Arshad, and in my opinion, it is fully adequate, in scope and quality, for the degree of M. Phil leading to Ph.D. Furthermore, it is hereby approved for submission for review and thesis defense. Supervisor: _____________________ Name: Dr. Muhammad Sajjad Mirza Date: 27 December, 2016 Place: NIBGE, Faisalabad Co-Supervisor: __________________ Name: Dr. Shaheen Asad Date: 27 December, 2016 Place: NIBGE, Faisalabad Head, Department of Biotechnology: ___________________ Name: Dr. Shahid Mansoor (S.I) Date: 27 December, 2016 Place: NIBGE, Faisalabad Bacterial Diversity in the Rhizosphere of AVP1 Transgenic Cotton (Gossypium hirsutum L.) and Wheat (Triticum aestivum L.) Muhammad Arshad Submitted in partial fulfillment of the requirements for the degree of Ph.D. 2016 Department of Biotechnology Pakistan Institute of Engineering and Applied Sciences Nilore-45650 Islamabad, Pakistan Dedications To My Parents & My innocent kids Muhammad and Anaya ii Acknowledgements Nothing is deserving of worship except “ALMIGHTY ALLAH”, all praises for Him, Who is the entire source of all knowledge and wisdom endowed to mankind. He guides the way and gives me courage to complete this work. I offer my humblest gratitude from deep sense of heart to the Holy Prophet, MUHAMMAD (Peace be Upon Him) Who is, forever source of guidance and knowledge for humanity. I am very grateful to my PhD supervisor Dr. Muhammad Sajjad Mirza, Deputy Chief Scientist, NIBGE Faisalabad for his professional and technical guidance, scientific discussions and suggestions, keen interest in completion of this task and moral support during whole period of research and compilation of thesis. I also pay thanks to my foreign supervisor Professor Dr. Johan Leaveau at Plant Pathology Department University of California Davis CA, USA for his kind and technical support and valuable contribution during my visit to the host lab for six month fellowship. I will appreciate Mr. Gurdeep Rastogi, my senior colleague and Mr. Jan Tech, lab in charge at Pathology Lab, University of California Davis CA, USA. I would also like to appreciate and acknowledge the efforts of Dr. Shahid Mansoor (S.I.), Director (NIBGE), and Dr. Suhail Hameed (Exe. Director NIBGE) for maintaining the honor of this institute among other research organizations of Pakistan. I would like to acknowledge Dr. Shaheen Asad (co-superviser) and Dr. Nasir A saeed Principle Scientists at NIBGE, Faisalabad, for providing all plant material. I would like to appreciate Mr. Muhammad Arshad Senior Scientist at NIBGE and Mr. Masood Anwar for their cooperation during this course of study. I am also indebted to my lab colleagues Dr. Muther Mansoor Qaisrani, Dr. Muhammad Tahir and Mr. Ahmad Zaheer for their help in learning research techniques and theoretical discussions. I am also thankful to Mr. Muhammad Ahmad and Muhammad Imran technicians at Microbial Ecology Lab, NIBGE, Faisalabad, for the kind help in conducting lab and field experiments. The help from Dr. Farooq iii Latif (DCS) and Dr. Ghulam Rasul (PS), at NIBGE, Faisalabad for analysis of organic acids and phytohormones on HPLC is thankfully acknowledged. Special thanks are due to my parents who waited a long time. I am especially thankful to my wife Aisha Arshad who suffered my long absence at home brought up my beloved kids Muhammad and Anaya with full care and provided me the spiritual and moral support during this long period of study, research work and in thesis writing. Many friends have helped me stay sane through these difficult years. Particularly, I am thankful to Dr. Atif Iqbal, Dr. Asif Habib Dr. Ikram Anwar and Sohail Mehmood Kareemi. Their support and care helped me to overcome setbacks and stay focused on my graduate study. I greatly value their friendship and I deeply appreciate their belief in me. I have no words to pay sincerest thanks to my friends for their help, encouragement and great friendship that made easier for me to overcome difficulties in all hard times. At the end, I would like to acknowledge Higher Education Commission, Pakistan, for providing me funds for my doctoral research in Pakistan and University of California, Davis CA USA. Without this financial support I might not be able to focus on my research. Muhammad Arshad iv Declaration of Originality I hereby declare that the work accomplished in this thesis is the result of my own research carried out in Soil & Environmental Biotechnology Division (NIBGE). This thesis has not been published previously nor does it contain any material from the published resources that can be considered as the violation of international copyright law. Furthermore, I also declare that I am aware of the terms ‘copyright’ and ‘plagiarism’, and if any copyright violation was found out in this work I will be held responsible of the consequences of any such violation. __________________ (Muhammad Arshad) 27 December, 2016 NIBGE, Faisalabad. v Copyrights Statement The entire contents of this thesis entitled Bacterial Diversity in the Rhizosphere of AVP1 Transgenic Cotton (Gossypium hirsutum L.) and Wheat (Triticum aestivum L.) by Muhammad Arshad are an intellectual property of Pakistan Institute of Engineering & Applied Sciences (PIEAS). No portion of the thesis should be reproduced without obtaining explicit permission from PIEAS. vi Table of Contents Dedications .................................................................................................................... ii Acknowledgements ...................................................................................................... iii Declaration of Originality .............................................................................................. v Copyrights Statement .................................................................................................... vi Table of Contents ......................................................................................................... vii List of Figures ............................................................................................................... xi List of Tables .............................................................................................................. xiv Abstract ...................................................................................................................... xvii List of Publications and Patents .................................................................................. xix List of Abbreviations and Symbols.............................................................................. xx 1. Introduction ................................................................................................................ 1 1.1 Genetically Modified Crops .................................................................................. 1 1.2 Use of AVP1 Gene to Develop Transgenic Plants ................................................ 2 1.3 Bacterial Diversity in Rhizosphere of Genetically Modified Plants ..................... 4 1.4 Plant Growth Promoting Rhizobacteria (PGPR) .................................................. 5 1.4.1 Mode of Action of PGPR .......................................................................... 5 1.4.2 Nitrogen Fixation ...................................................................................... 6 1.4.3 Biological Nitrogen Fixation (BNF) ......................................................... 8 1.4.4 Diversity of Diazotrophic Bacteria ........................................................... 9 1.4.5 The Domain Archea .................................................................................. 9 1.4.6 Phosphorus Mineralization by Microbes for Plant Growth Promotion .. 11 1.4.7 Phytohormone Production by PGPR for Plant Growth Promotion ........ 12 1.4.8 PGPR as Biofertilizers ...........................................................................
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