Title Page
Contents
ABSTRACT 10
Ⅰ. Introduction 13
1.1. The NSD gene family and its function 13
1.2. Heterochromatin formation and epigenetic regulations 16
1.3. Gene amplification in Drosophila 21
Ⅱ. Materials and Methods 25
2.1. Drosophila strains 25
2.2. Transcriptome analysis 25
2.3. Generation of NSD deletion lines on various mini-white transgene backgrounds 26
2.4. Drosophila genotyping by PCR 27
2.5. Quantitative assessment of Drosophila eye pigment 28
2.6. RNA extraction and reverse transcription-quantitative PCR (RT-qPCR) 28
2.7. Statistics 30
Ⅲ. Results 31
3.1. Effects of NSD deletion on gene silencing in Drosophila 31
3.1.1. Transcriptome analysis of the adult ovary with NSD deletion 31
3.1.2. Generation of heterozygous and homozygous NSD deletion mutant flies on various mini-white transgene backgrounds 36
3.1.3. The differential effects of NSD deletion on expression of the mini- white gene localized at various chromosomal sites 47
3.2. Effects of NSD deletion on gene amplification in Drosophila 57
3.2.1. The effect of NSD deletion on transcriptome analysis in the DAFCs region in Drosophila 57
3.2.2. The effect of NSD deletion on the DNA amplification and transcription of the genes in the DAFCs regions from the Drosophila adult ovary 61
Ⅳ. Discussion 64
References 68
Abstract (in Korean) 75
Table 1. Primer pairs for PCR genotyping 27
Table 2. Primer pairs for quantitative PCR (mRNA) 29
Table 3. Primer pairs for quantitative PCR (genomic DNA) 30
Table 4. Transcriptome raw data of w¹¹¹⁸ and the NSD null mutant fly. 33
Table 5. Confirmation of the transcriptome results NSD null mutant fly by qPCR. 33
Table 6. The relative levels of eye pigment of heterozygous as well as homozygous NSD deletion mutant flies in different mini-w transgene... 56
Table 7. Relative expression levels of genes located in the two DAFC regions, DAFC-62D and DAFC-66D, analyzed from the list of... 59
Figure 1. The NSD family members and their architecture with functional domains. 15
Figure 2. Position effect variegation in Drosophila. 20
Figure 3. DAFC regions in Drosophila. 24
Figure 4. Analysis of transcriptome data of the NSD null mutant fly (whole genes). 34
Figure 5. Analysis of the transcriptome data of the NSD null mutant fly. (Genes displaying more than two-fold differences) 35
Figure 6. Genomic and biochemical analysis of the NSD deletion fly used in this study. 38
Figure 7. Drosophila strains used in this study and their characteristics. 41
Figure 8. Generation of homozygous and heterozygous NSD deletion mutants on various mini-w transgene backgrounds by crossing or recombination. 44
Figure 9. Genomic PCR of homozygous and heterozygous NSD deletion flies for genotyping. 45
Figure 10. Crossing result of mini-w located on the X chromosome in Drosophila male eyes and the effect of NSD null mutation. 46
Figure 11. The dose effect of variegating transgenes, mini-w, on various location on the eye phenotype. 51
Figure 12. Effects of heterozygous and homozygous NSD-deletion mutations on PEV. 53
Figure 13. The statistical analysis represented in Figure 12 (B). 55
Figure 14. The relative mRNA levels of genes highly expressed during mid- oogenesis in the ovary of NSD null mutant virgin females. 60
Figure 15. The relative levels of amplificated DNAs in the DAFCs of the NSD null mutant ovaries. 63