Muhammad Arif Nadeem Saqib_Biochemistry_&_Molecular_Biology_2015_QAU_ISD

194 Pages·2015·3.15 MB·English

GENETIC ANALYSIS OF RARE EYE DISORDERS
IN PAKISTANI FAMILIES

By

MUHAMMAD ARIF NADEEM SAQIB

Department of Biochemistry
Faculty of Biological Sciences
Quaid-i-Azam University
Islamabad, Pakistan
2015
GENETIC ANALYSIS OF RARE EYE DISORDERS
IN PAKISTANI FAMILIES

A thesis
Submitted in the partial fulfillment of the
Requirements for the degree of
DOCTOR OF PHILOSOPHY
In
BIOCHEMISTRY/MOLECULAR BIOLOGY
By

MUHAMMAD ARIF NADEEM SAQIB

Department of Biochemistry
Faculty of Biological Sciences
Quaid-i-Azam University
Islamabad, Pakistan
2015
CERTIFICATE

A thesis submitted in the partial fulfillment of the requirements
for the degree of the Doctor of Philosophy. We accept this
dissertation as conforming to the required standard.

1 _________________ 2________________
Dr. Muhammad Ansar External Examiner
(Supervisor)

3 ____________________
Dr. Muhammad Ansar
(Chairperson)

Dated: February 09, 2015
DECLARATION

I hereby declare that the work presented in the following thesis is my own
efforts and that the thesis is my own composition. No part of the thesis
has been previously presented for any other degree.

Muhammad Arif Nadeem Saqib
Dedicated
To

Muhammad Romman Khan
(Late)
My Son, who died during the journey of my PhD
ACKNOWLEDGEMENT
All praise to Allah Almighty, the most beneficent, the most merciful, Who
gave me strength and enabled me to undertake and execute this research task.
Countless salutations upon the Holy Prophet Hazrat Muhammad (Sallallaho Allaihe
Waalahe Wassalum), the city of knowledge for enlightening with the essence of faith
in Allah and guiding the mankind, the true path of life.
I would like to extend my appreciation to those people, who helped me in one
way or another to finish the thesis. It is a pleasure to convey my gratitude in my
humblest acknowledgment to those people who contributed to make the thesis.
I feel highly privileged in taking the opportunity to express my deep sense of
gratitude to the Dr. Muhammad Ansar, Chairperson and Associate Professor,
Department of Biochemistry, Faculty of Biological Sciences, QAU, Islamabad, for his
scholastic guidance and valuable suggestions throughout the study and presentation
of this dissertation. I am thankful to him for his inspiration, reassurance, counseling
from time to time and specially a friendly environment.
I am thankful to Dr. Carlo Rivolta, Assistant Professor, Department of
Medical Genetics, University of Lausanne, Lausanne, Switzerland, who provided me
opportunity to work in his lab under International Research Support Initiative
Program of Higher Education Commission, Pakistan. He remained very kind and
boosted my scientific skills.
I am obliged to Dr. Bushra Mirza, Ex-Chairperson, Department of
Biochemistry, Faculty of Biological Sciences, Quaid-i-Azam University (QAU),
Islamabad, for extending the research facilities of the department to accomplish this
work.
I am also obliged to my all teachers who contributed a lot in my studies,
especially to Dr. Azra Khanum, Prof of Biochemistry and Sir Sharif Amjad, Science
Teacher, Government High School, Ghangwal.
Special thanks to Dr. Huma Qureshi, Executive Director, PMRC, Islamabad,
Dr. Arif Munir, Principal Research Officer, PMRC, Islamabad and Ibrar Rafique,
Research Officer, PMRC, Islamabad who provided a lot of scientific insights and help
in learning the research writing.
Heartiest and sincerest acknowledgement also goes to Higher
Education Commission of Pakistan for granting me Indigenous as well as IRSIP (for
foreign research support) scholarship, without which this achievement was not
possible. We would like to particularly thank all members of the families that
participated in this study for their invaluable participation and cooperation
I am also thankful to my lab fellows Dr. Muzamal Ahmad Khan, Falak Sher,
Rani Sara, Noureen Aslam, Waleed Ahmed Khan, Naeem Ashraf, Madiha Amin,
Neelum Shah, Jamil Iqbal, Sumaira Anjum, Nikhat Siddique, Muhammad Nawaz,
Farah Fatima, Rabia, Mehwish, Benish, Jawad Akhtar, Kinza Makhdoom, Sundus
Sajid, Khalid Khan, Afeefa Jarral, Noor Han, Zoma Khan, Zainab Akram, Riffat
and Memoona Rasheed, who assist me in various ways during my research work
Thanks to all the colleagues in Carlo’s lab, especially Konstantinos
Nikopoulos, Giulia Venturini, Hanna Koskiniemikundig, Alessandro DI Gioia,
Paola Benaglio, Pietro Farinelli, Rosanna Pescini Gobert, Nicola Bedoni, and
Beryl Royer-Bertrand for their support and company.
Thanks to my friends Ehsan Ullah, Ahmad Waqas, Muhammad Umair,
Younus Khan, Haroon Khan, Safdar Abbas, Waqas Kayani for their company,
encouragement, concern, laughters stories and for taking their time to listen to
everything that I had to say. Special thanks to Dr. Rizwan Alam, Dr. Khalid Khan,
Dr. Muhammad Jadoon Khan for their company and time to visit different cities of
Europe. It was really pleasant to visit them and see the beauty of Europe.
I would like to acknowledge the clerical staff of the Department of
Biochemistry especially to Mr. Tariq, Fayaz, Saeed and other staff members for their
support and help. I would also like to thank Mr Saeed Ahmad Shahid, Hafiz Saqib,
Junaid.
Last but not least all goes to my mother (late), who enlightened me and whose
prayers still remained in search of my successes. She will remain always alive in my
heart. My loving father, Hafiz Abdul Hameed, who remained a guide through my
studies and provided me his best, my elder brother, Hafiz Ahmad Tariq, who gave me
the strength and courage and provided all opportunities to achieve this task, my
younger brother, Hafiz Atif Naveed, who is always a source of joy and provide his
best for my leisure times, my all sisters who deserve special mention for their
inseparable support and prayers throughout my life. It wouldn’t have been this
bearable if I didn’t have these in my life. Thank you for your unconditional support
with my studies. Thank you for giving me a chance to prove and improve myself
through all walks of my life. Bundle of thanks to my uncle, Abdul Waheed Zafar,
and Mian Mehboob ur Rasool (Late), brother in laws (Bhai Ihsan, Bhai Akhtar,
Qamar Bhai, Bhai Sohail, Bhai Waqas, Mustansar, Sohaib and Yasir Din) and
cousins for providing their continuous support.
Special thanks to my beloved wife for her love, devotion and constantly
reminding me to take care of myself. Special thanks to Khans (my sons: Muhammad
Somaan Khan and Muhammad Affan Khan and my nephews: Muhammad Sinnan
Khan and Muhammad Siban Khan) who provides joy by their naughtiest actions.
Finally, I would like to thank everybody who was important to the successful
completion of the thesis, as well as expressing my apology that I could not mention
personally one by one.
Muhammad Arif Nadeem Saqib
______________________________________________________________Contents
TABLE OF CONTENTS
Pages
List of Figures ii
List of Tables iv
List of Abbreviations vi
Abstract xi

CHAPTER 1 1
1.0 Introduction 2
1.1 Rare Eye Disorders 2
1.2 Inherited Retinal Disorders 4
1.3 Rod Associated Retinal Disorders 4
1.3.1 Congenital Stationary Night Blindness 7
1.3.1.1 Genetics of CSNB 7
1.3.1.1.1 Genes Responsible for Complete CSNB 8
1.3.1.1.2 Genes Responsible for Incomplete CSNB 9
1.3.2 Retinitis Pigmentosa 10
1.3.2.1 Genetics of Retinitis Pigmentosa 11
1.3.2.1.1 RP Genes of Phototransduction Cascade 11
1.3.2.1.2 RP Genes of Splicing and Transcription 12
1.3.2.1.3 Ciliary Gene 13
1.4 Cone Associated Retinal Disorders 14
____________________________________________________________________
Genetic Analysis of Rare Eye Disorders in Pakistani Families
______________________________________________________________Contents
1.4.1 Achromatopsia 14
1.4.1.1 Genetics of Achromatopsia 15
1.4.2 Cone and Cone Rod Dystrophies 17
1.4.2.1 Genetics of Cone and Cone Rod Dystrophies 17
1.5 Syndromic Retinal Dystrophies 19
1.5.1 Bardet-Biedl Syndrome 19
1.5.2 Usher Syndrome 20
1.5.3 Cohen Syndrome 20
Genetic Analysis of Retinal Dystrophies in Pakistani
1.6 21
Families
1.7 Objectives 24
CHAPTER 2 25
2.0 Materials and Methods 26
2.1 Families Studied 26
2.1.1 Clinical and Ophthalmologic Examination 26
2.2 Blood Sampling 27
2.3 Extraction of Genomic DNA from Human Blood 27
2.3.1 DNA Extraction by Organic Method 27
2.3.1.1 Composition of Solutions 28
2.3.2 DNA Extraction by Commercially Available Kit 28
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Genetic Analysis of Rare Eye Disorders in Pakistani Families