Title page
ACKNOWLEDGEMENT
Contents
ABBREVIATIONS 11
CHAPTER 1. INTRODUCTION 12
1. Infertility 13
1-1. Male infertility 13
1-1-1. Unexplained Male Infertility 14
1-1-2. Azoospermia 14
1-2. Single Nucleotide Polymorphism 15
1-3. SNPs and Disease Diagnosis 16
CHAPTER 2. ASSOCIATION STUDY OF SINGLE NUCLEOTIDE POLYMORPHISM WITH MALE INFERTILITY 17
2-1. ABSTRACT 18
2-2. INTRODUCTION 19
2-3. MATERIALS AND METHODS 22
2-3-1. Study population 22
2-3-2. Cytogenetic and Y chromosome microdeletion assay 23
2-3-3. PCR and restriction fragment length polymorphism (RFLP) for INSL3 G178A polymorphism 24
2-3-4. PCR Length Polymorphism (LP) for 4a4b Polymorphism 24
2-3-5. Pyrosequencing assay of eNOS -786T〉C, 894G〉T Polymorphisms 24
2-3-6. Statistical analysis 25
2-4. RESULTS 27
2-5. DISCUSSION 30
REFERENCES 44
Table 1. Cause of male infertility 32
Table 2. Clues to the Diagnosis of Male Infertility 33
Table 3. Semen analysis for male infertility 34
Table 4. The primer sequences for pyrosequencing assay for -786T〉C and 894G〉T sites of endothelial nitric oxide synthase (eNOS) 35
Table 5. List of insulin-like factor 3 (INSL3) SNPs and Mutations 36
Table 6. Statistical analysis of INSL3 G178A genotype from infertile and fertile males 37
Table 7. The frequency of INSL3 G178A genotype according to the subtype of infertility 38
Table 8. Sperm count, motility, vitality, morphology according to genotype of INSL3 in the ejaculate semen group 39
Table 9. Genetic frequency of endothelial nitric oxide synthase (eNOS) -786T〉C, 4a4b, 894G〉T polymorphisms in infertile and fertile men. 40
Table 10. Sperm count, motility, and morphology according to genotype of eNOS in the ejaculate semen groups 41
Fig. 1. PCR-RFLP results according to the INSL3 G178A genotype from fertile males (Controls) and infertile males (Patients). Homozygous 178AA has one restriction site and generated two fragments of 195 bp and 26 bp (lanes 1 and 4). Heterozygous 178GA types have four fragments of 195 bp, 121 bp,... 43