Wholegenome shotgun assembly and analysis of the genome of Fugu rubripes pufferfish Presented by Cha

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Whole-genome shotgun assembly and analysis of the
genome of Fugu rubripes (pufferfish) Presented
by Charu Gupta Kumar

• Aparicio et al and Brenner S. Whole-genome
  shotgun assembly and analysis of the genome of
  Fugu rubripes. Science. 2002 Aug
  23297(5585)1301-10.

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Classification of bony vertebrates.

• Euteleostomi (bony vertebrates)
• Actinopterygii (ray-finned fishes)
• Actinopteri
• Chondrostei
• Neopterygii ?Teleosts ? ??Tetraodonts
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  ?Fugu Rubripes
• Polypteriformes
• Polypteridae (bichirs)
• Sarcopterygii (lobe-finned fishes)
• Coelacanthimorpha
• Coelacanthiformes
• Dipnoi (lungfishes)
• Ceratodontimorpha
• Tetrapoda (tetrapods)
• Amniota (amniotes) ??Mammalia??Humans
• Amphibia (amphibians)

450 million years since their last common ancestor
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Fugu Genome Human Genome

• 365 Mbp genome
• 31,059 predicted gene loci
• 33,609 predicted peptides
• Intron size is 79 bp modal, with 75 lt 425bp
• 1/6 of genome is repetitive elements
• Fugu has reduced intergenic regions
• Number of introns is comparable in both Fugu and
  Human

• 2.91 Gbp genome
• 29,181 predicted gene loci
• 31,780 nonredundant peptides
• Intron size is 87 bp modal, with
• 75 lt 2605bp
• 43 of genome is repetitive elements

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Preliminary Annotation

• 33,609 predicted Fugu peptides
• Gene containing fraction of genome is a mere 108
  Mb of the euchromatic 320Mb
• Intron-exon structure of most genes is preserved
  between Fugu and Human
• Conservation of synteny between Fugu and Human.
• Allows one to identify chromosomal elements from
  the common ancestor
• 75 of Fugu loci have Tblastx hits against human
  genome

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Identification of novel putative gene loci in Hs.
Fugu Proteins
Blast against Ensembl Hs peptides
8,761 Fugu proteins have hits gt 10-3 (27,779
have hits lt 10-3)
Tblastn against masked Hs genomic sequence
1800 match
Used these as input to build Hs genes through an
EnsEMBL Hs pipeline
1260 remain as apparent Novels
Filter for low-complexity peptides, search NCBI
again
961 Fugu proteins that are novel gene loci in
Humans
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Genome Analysis.

• RepeatMasker analysis showed 2.7 of genome
  matches interspersed repeats (LINEs, SINEs, MIR,
  etc)
• Total number of introns about the same in Fugu
  and Humans.
• Majority of Fugu gene loci are scaled in
  proportion to the compact genome size.
• Observed intronless Fugu orthologs in Humans.
  But some genes were 1.3X or greater in size
  compared to Human

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Distribution of ratios for gene locus sizes of
putative Fugu-human orthologous pairs.
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Conservation of synteny.

• Orthologous gene loci are linked in two species,
  regardless of gene order or presence of
  intervening genes.

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Location of conserved Fugu segments in the human
genome.
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Duplications in Fugu Genome

• Compared Fugu to itself using windows of 1 kb and
  500bp.
• 0.15 and 1.3 respectively of the Fugu genome
  contained duplicated segments.
• In Humans, 5 of the genome was duplicated in
  segments of gt 1 kb.
• More evidence comes from existence of ancient
  paralogous segments. When occur after speciation
  event, called in-paralogs.

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Comparison of Fugu and Human predicted proteomes.

• 3/4 of predicted human proteins have a strong
  match to Fugu.
• Remaining 1/4 (8109) appear to have no pufferfish
  homologs.
• Reciprocally, 6000 (19) Fugu predicted proteins
  lack significant homology in human.
• A significant part of this 8109 set could
  represent
• evolution of proteins between two vertebrates so
  that they are no longer mutually recognizable at
  the sequence level, or
• Loss of genes common to other vertebrates in
  Fugu, or
• Gain of sequences specific to tetrapod or
  mammalian orders, or
• Erroneous human gene predictions

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Distribution of protein similarities between Fugu
and Human proteomes.
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In conclusion

• Compact Fugu genome, and low abundance of
  repeats. Easy to characterize regulatory elements
  in noncoding sequence
• More similarities than differences between Fugu
  and Human
• Rapid evolution of proteins may account for most
  of the observable differences.
• Often gene order and orientation preserved in
  vertebrates.
• Comparative Analysis with Fugu genome powerful
  for discovering vertebrate homologs, and
  discovering synteny.