Object 4-1: Supplementary information for Chapter 4

Supplementary Figure S1. The phylogenetic tree constructed using 21 cyanobacteria with complete genome assemblies and quantified ploidy level from Zborowsky & Lindell, 2019. The 16S sequence was extracted from their genomes and trimmed to ensure the perfect overlap. Bootstrapping was performed 1,000 times for assessing the confidence of branching determination. Branching points within Prochlorococcus sp. has been significantly higher than Synechococcus sp. Atribacterota Balneolota Candidatus Cloacimonadota Candidatus Dadabacteria Candidatus Omnitrophota Chlorobiota Chrysiogenota Coprothermobacterota Dictyoglomota Kiritimatiellota Chlamydiota Bacillota Fusobacteriota Verrucomicrobiota Synergistota Spirochaetota Lentisphaerota Candidatus Binatota Deferribacterota Fibrobacterota Bdellovibrionota Thermodesulfobacteriota Elusimicrobiota Deinococcota Chloroflexota Bacteroidota Candidatus Kapabacteria Candidatus Tectomicrobia Mycoplasmatota Pseudomonadota Campylobacterota Candidatus Latescibacteria Nitrospirota Rhodothermota Acidobacteriota Gemmatimonadota Thermotogota Actinomycetota Armatimonadota Planctomycetota Ignavibacteriota Myxococcota Aquificota Cyanobacteriota Candidatus Poribacteria Abditibacteriota Caldisericota Calditrichota Candidatus Bipolaricaulota Candidatus Eremiobacterota Candidatus Fervidibacteria Candidatus Melainabacteria Nitrospinota Thermomicrobiota candidate division NC10 0.00 0.25 0.50 0.75 1.00 Proportion Phylum 1 2 3 Species Genus Family Order Class 11111 22221 22111 11111 11111 11111 11111 11111 11111 11111 11111 656133101 1010432 1710842 33321 44401294 44444 98423264307 2612541 53233 2116775 44411 96532 11111 117211132 2 , 183 735151476 13920863 11111 11111 4661844197 1715101010 367331 43333 8854241311 76433 22222 77511 11111 22232 8220941 88722 2016864 299311 96632560177 259521 33333 21111 11111 11111 116322 11111 11111 11111 11111 11111 Atribacterota Balneolota Candidatus Cloacimonadota Candidatus Dadabacteria Candidatus Omnitrophota Chlorobiota Chrysiogenota Coprothermobacterota Dictyoglomota Kiritimatiellota Chlamydiota Bacillota Fusobacteriota Verrucomicrobiota Synergistota Spirochaetota Lentisphaerota Candidatus Binatota Deferribacterota Fibrobacterota Bdellovibrionota Thermodesulfobacteriota Elusimicrobiota Deinococcota Chloroflexota Bacteroidota Candidatus Kapabacteria Candidatus Tectomicrobia Mycoplasmatota Pseudomonadota Campylobacterota Candidatus Latescibacteria Nitrospirota Rhodothermota Acidobacteriota Gemmatimonadota Thermotogota Actinomycetota Armatimonadota Planctomycetota Ignavibacteriota Myxococcota Aquificota Cyanobacteriota Candidatus Poribacteria Abditibacteriota Caldisericota Calditrichota Candidatus Bipolaricaulota Candidatus Eremiobacterota Candidatus Fervidibacteria Candidatus Melainabacteria Nitrospinota Thermomicrobiota candidate division NC10 0.00 0.25 0.50 0.75 1.00 Proportion Phylum 1 2 3 Supplementary Figure S2. The phylogenetic tree constructed using 21 cyanobacteria with complete genome assemblies and quantified ploidy level from Zborowsky & Lindell, 2019. The 16S sequence was extracted from their genomes and trimmed to ensure the perfect overlap. Bootstrapping was performed 1,000 times for assessing the confidence of branching determination. Branching points within Prochlorococcus sp. has been significantly higher than Synechococcus sp. Synechococcus CC9605 Synechococcus WH8109 Synechococcus RS9917 Synechococcus BL107 Synechococcus CC9902 Synechococcus CC9311 Synechococcus WH7803 Synechococcus WH7805 Synechococcus RCC307 Prochlorococcus MIT9313 Prochlorococcus NATL2A Prochlorococcus SS120 Prochlorococcus MIT0604 Prochlorococcus MIT9215 Prochlorococcus MIT9312 Synechococcus RS9916 Synechococcus WH8102 Prochlorococcus MED4 Prochlorococcus MIT9515 38 100 78 53 85 40 32 71 91 90 44 51 48 95 31 38 Tree scale: 1 Supplementary Figure S3. The phylogenetic tree constructed using 8 Prochlorococcus sp., with complete genome assemblies and quantified ploidy level from Zborowsky & Lindell, 2019. The 16S sequence was extracted from their genomes and trimmed to ensure the perfect overlap. Bootstrapping was performed 1,000 times for assessing the confidence of branching determination. Species with orange font are monoploids, and species with blue font are polyploids. Prochlorococcus MIT9313 Prochlorococcus NATL2A Prochlorococcus SS120 Prochlorococcus MIT0604 Prochlorococcus MIT9215 Prochlorococcus MIT9312 Prochlorococcus MED4 Prochlorococcus MIT9515 81 69 95 76 95 Tree scale: 0.01 Supplementary Figure S4. Relationship between pairwise Euclidean distance and evolutionary distance. Pairwise Euclidean distance was calculated using the coordinates of two randomly selected samples from 3D space formed by first three principal components using global genomic architecture. Evolutionary distance was calculated based on the phylogenetic tree IQTREE v3 with 3,000 samples from GTDB set. Different colors reflect the evolutionary relationship with warmer color representing closer relatives. For example, orange points represent sample pairs from the same species, and dark blue points represent sample pairs from the same kingdom. Supplementary Figure S5. Distributions of proportion of genes located on the lagging strand from three different groups. Overall, among three groups there were similar number of genes located at the lagging strand. However, group 2 and group 3 bacteria have narrower distributions of the percentage of genes on lagging strand, indicating genomic recombination promoted the inversion in a random way. Two-sample Kolmogorov-Smirnov tests were conducted between two groups to examine whether samples from two groups are from the same distribution. Group 2 and group 3 significantly differentiate from group 1, while the null hypothesis of the same distribution cannot be rejected comparing group 2 and group 3 distribution p = 1.761e−06 p = 0.06062 p = 4.941e−09 0.00 0.25 0.50 0.75 1.00 1.25 123 Group % of genes on lagging strand Supplementary Figure S6. The heatmap shows the orientational bias of different COG functional categories that are located on the lagging strand through the odds ratio obtained from Fisher’s exact tests. COG entries that were mapped to more than one functional category were tolerated and plotted here. Most COG categories in group 1 have orientational bias on either leading strand (yellow) or lagging strand (blue). However, most COG categories in group 2 and group 3 do not show significant orientational bias (grey). Benjamini–Hochberg correction was performed on all COG categories to control the false discovery rate. 1 2 3 KV XT HT HQ KN Z QV GT MN MT OT JE OJ PE JT LX DKO KE PT FR W QR JL KG MO HR JO KT MU GM NW P X K R S T HJ E ER O V EQ EG EP J U C N F G L QC TR D PR TG FM JR DE DO JHO DL MI DM KJ OK CO B FT −3 −2 −1 0 1 2 3 COG functional category log₂(odds ratio) 1 2 3 COG category Full name ARNA processing and modification BChromatin structure and dynamics CEnergy production and conversion DCell cycle control, cell division, chromosome partitioning EAmino acid transport and metabolism FNucleotide transport and metabolism GCarbohydrate transport and metabolism HCoenzyme transport and metabolism ILipid transport and metabolism JTranslation, ribosomal structure and biogenesis KTranscription LReplication, recombination and repair MCell wall/membrane/envelope biogenesis NCell motility OPosttranslational modification, protein turnover, chaperones PInorganic ion transport and metabolism QSecondary metabolites biosynthesis, transport and catabolism RGeneral function prediction only SFunction unknown TSignal transduction mechanisms UIntracellular trafficking, secretion, and vesicular transport VDefense mechanisms WExtracellular structures YNuclear structure ZCytoskeleton 1 2 3 KV XT HT HQ KN Z QV GT MN MT OT JE OJ PE JT LX DKO KE PT FR W QR JL KG MO HR JO KT MU GM NW P X K R S T HJ E ER O V EQ EG EP J U C N F G L QC TR D PR TG FM JR DE DO JHO DL MI DM KJ OK CO B FT −3 −2 −1 0 1 2 3