Title Page
Contents
ABSTRACT 10
INTRODUCTION 13
REFERENCES 16
Chapter 1. Bacterial diversity of Lebbeus groenlandicus and Pandalopsis japonica by 16S rRNA gene amplicon sequencing 18
1.1. Abstract 18
1.2. Introduction 20
1.3. Materials and Methods 23
1.3.1. Sample collection and DNA extraction 23
1.3.2. Barcode PCR and NGS sequencing 23
1.3.3. Bioinformatic analysis of reads 27
1.4. Results and discussion 28
1.5. Conclusion 35
1.6. References 36
Chapter 2. Isolation and genomic analysis of M13 and M17, novel psychrophilic bacteria, isolated from Lebbeus groenlandicus 38
2.1. Abstract 38
2.2. Introduction 40
2.3. Materials and Methods 42
2.3.1. Cultivation and isolation 42
2.3.2. Colony PCR 42
2.3.3. Phylogenetic analysis 43
2.3.4. Morphological and biochemical tests 43
2.3.5. Polar lipids 44
2.3.6. Whole genome sequencing 45
2.4. Results and discussion 46
2.5. Conclusion 64
2.6. References 65
국문초록 67
Table 1. The list of barcoded PCR primer sequences used in this study 25
Table 2. 16S rRNA sequence similarity of strain M13 and other strains showed ten closest strains on the ez-biocloud server 47
Table 3. 16S rRNA sequence similarity of strain M17 and other strains showed ten closest strains on the ez-biocloud server 47
Table 4. Phenotypic characteristics that differentiate strain 52
Table 5. Assembly results of M13 and M17 56
Table 6. BUSCOs results of M13 and M17 56
Table 7. Detailed information of a prophage detected in the genome of strain M13 60
Table 8. Detailed information of a prophage detected in thegenome of strain M17 61
Table 9. Identification and prediction of M13 genes involved in the biosynthesis of secondary metabolites using antiSMASH 62
Table 10. Identification and prediction of M17 genes involved in the biosynthesis of secondary metabolites using antiSMASH 62
Table 11. Results of ANI (average nucleotide identity) analysis between the strain M13 and M17 63
Table 12. Results of AAI (average amino acid identity) analysis between the strain M13 and M17 63
Figure 1. Unweighted UniFrac PCoA plot of shrimp microbiome (G: Lebbeus groenlandicus, J: Pandalopsis japonica) 29
Figure 2. Diversity of shrimp microbiome at class level (G: Lebbeus groenlandicus, J: Pandalopsis japonica) 30
Figure 3. Diversity of shrimp microbiome at genus level (G: Lebbeus groenlandicus, J: Pandalopsis japonica) 31
Figure 4. Comparison of diversity index by species (G: Lebbeus groenlandicus, J: Pandalopsis japonica) 33
Figure 5. Comparison of diversity index by species and organs (G: Lebbeus groenlandicus, J: Pandalopsis japonica) 33
Figure 6. Comparison of mean values of taxa at the genus level by species and organs 34
Figure 7. Unweighted UniFrac PCoA plot of shrimp microbiome (by species and organs) 34
Figure 8. Phylogenetic tree of the candidates of novel species Numbers indicate bootstrap percentages from neighbour-joining after 1000 replication; values... 48
Figure 9. Phylogenetic tree showing the position of strain M13 and M17 Numbers indicate bootstrap percentages from neighbour-joining after 1000 replication; values... 49
Figure 10. Results of gram staining of strains M13 and M17 under an optical microscope (1,000X) 51
Figure 11. Two-dimensional TLC of the total polar lipids of strain M13 (left) and M17 (right) 53
Figure 12. Genome map of M13 and M17 genomic DNA 57
Figure 13. Subsystems of (A) M13 and (B) M17 genomic DNA annotated by Rapid Annotations using Subsystems Technology 58
Figure 14. Metabolic pathway of M13 and M17 using KEGG 59
Figure 15. Metabolism aromatic compounds pathway using KEGG red: the metabolic pathway of M17, blue: the metabolic pathways of M13 and M17 59
Figure 16. Location of prophage sequences in the M13 genome 60
Figure 17. Location of prophage sequences in the M17 genome 61
Figure 18. Secondary metabolite region of M13 62
Figure 19. Secondary metabolite region of M17 62