Genic regions of a large salamander genome contain long introns and novel genes

Abstract

Background: The basis of genome size variation remains an outstanding question because DNA sequence data are lacking for organisms with large genomes. Sixteen BAC clones from the Mexican axolotl (Ambystoma mexicanum: c-value = 32 x 10(9) bp) were isolated and sequenced to characterize the structure of genic regions.

Results: Annotation of genes within BACs showed that axolotl introns are on average 10x longer than orthologous vertebrate introns and they are predicted to contain more functional elements, including miRNAs and snoRNAs. Loci were discovered within BACs for two novel EST transcripts that are differentially expressed during spinal cord regeneration and skin metamorphosis. Unexpectedly, a third novel gene was also discovered while manually annotating BACs. Analysis of human-axolotl protein-coding sequences suggests there are 2% more lineage specific genes in the axolotl genome than the human genome, but the great majority (86%) of genes between axolotl and human are predicted to be 1:1 orthologs. Considering that axolotl genes are on average 5x larger than human genes, the genic component of the salamander genome is estimated to be incredibly large, approximately 2.8 gigabases!

Conclusion: This study shows that a large salamander genome has a correspondingly large genic component, primarily because genes have incredibly long introns. These intronic sequences may harbor novel coding and non-coding sequences that regulate biological processes that are unique to salamanders.

Figure 1

Comparison of intron lengths among the axolotl (AM), human (HS), chicken (GG), and frog Xenopus tropicalis (XT) for cleavage and polyadenylation specific factor 5 (NUDT21) and 3-hydroxy-3-methylglutaryl-Coenzyme A reductase (HMGCR). One exon of HMGCR could not be identified in the X tropicalis genome.

Figure 2

Structure of two A. mexicanum miRNAs (AMmiRNA16, AMmiRNA23) that were predicted from axolotl introns. The red bases indicated positions where the predicted miRNA sequences show complete identify to small RNAs isolated from regenerating limbs.

Figure 3

Intron-exon structure of a novel axolotl salamander gene (AxNovel_3) discovered within BAC H3_4A11. Intron/Exon boundaries are represented by vertical black bars. The predicted coding sequence for AxNovel_3 is shaded in blue. Red figures join the relative locations of sequences in the transcript and genomic sequence.