Lukacs et al. BMC Genomics 2010, 11:154 http://www.biomedcentral.com/1471-2164/11/154 Sasa-UGA The Sasa-UGA locus is inserted in the IB region between the genes for SLC39A and RING1. Multiple salmon ESTs matched this locus [Genbank: CB498868 and CA043257] and assembled together (GRASP cluster 280267) they provide a full-length Sasa-UGA cDNA sequence with 81% sequence identity towards rainbow trout UGA [Genbank: AAP04358]. Sasa-UHA The two Sasa UHA genes identified in 184H23 were named UHA1 and UHA2 and both loci have matching ESTs. The duplicated UHA1 and UHA2 loci both showed more than 93% identity towards ESTs previously named UHA and UHB by Miller et al [15]. A distinct difference between UHA1 and UHA2 is a three nucleo- tide insertion (an extra amino acid) in the a2-domain in UHA2. UHA1 and UHA2 are oriented in the same direction and show high similarity towards each other, but approximately 40-50% identity towards other U-linage molecules. Two different ESTs matching the UHA1 locus were identified, and they are most likely splice variants. UHA1*0101 [Genbank: EG787974] con- tains 10 exons, while UHA1*0201 [Genbank: DW548896] contains 7 exons, with the last exon containing only a stop-codon. UHA1*0201 contains an inserted exon con- sisting of 32 bp after the a3 domain (exon4), which con- tributes to an earlier stop in the CDS compared to (UHA1*0101). The nomenclature used for these splice variants is only temporary, awaiting a nomenclature debate. S-lineage The SAA locus identified in 114L13 showed 96% aa identity to Onmy-SAA*0101 [Genbank: AAB57877], and we found a matching Atlantic salmon EST [Genbank: DY713846]. As mentioned above, the salmonid SAA sequence shows very little similarity towards other U-lineage genes, with for instance approx 30% identity to Sasa-UBA alleles. Sasa-SAA*0101 is a compact class I gene with small intron sizes compared to the other MHC class I genes. Sasa-SAA*0101 contains the same characteristics as Onmy-SAA*0101 [16] with six exons and five introns, and an incapacitated transposable ele- ment in intron 3. Z-lineage We found 4 Z-lineage genes, one in the extended IA region and three in the extended IB region and this is in accordance with Miller et al [15] who identified four different ZE exon 2 sequences. The previously reported Atlantic salmon Sasa-ZE*0101 sequence [Genbank: DQ099914] shows 91% sequence identity towards the IA linked ZAA locus, and most likely emerged from this locus. The ZAA sequence in the 129P21 BAC is named Page 7 of 17 Sasa-ZAA*0201, while the previously reported sequence Sasa-ZE*0101 should be renamed Sasa-ZAA*0101. We have also identified matching EST clones for ZBA [Genbank: DY730127 and EG827413] and ZCA [Genbank: DY740683 and DW559009] making these bona-fide genes, while ZDA lacks the leader peptide suggestive of a pseudo- gene. ZBA*0101 and ZCA*0101 show 96% sequence identity over 918 bp, while the remaining 3’ sequence shows little sequence identity. Comparing the separate exons of all the Z-lineage genes (ZAA, ZBA, ZCA, ZDAψ), the highest sequence identity overall was found in exon2 and exon3, ranging from 89-96% identity for exon2, 84-92% identity for exon3, while exon4 showed 69-86% identity (data not shown). Comparing the different exons of ZBA*0101 and ZCA*0101 we found 95%, 92% and 86% identity in exon2, exon3 and exon4 respectively. They are all orien- tated in a head to tail fashion. The exon-intron organization of the four Z-lineage MHC class I genes are quite different with 5, 6, 4 and 4 exons for ZAA, ZBA, ZCA and ZDAψ respectively. The connecting peptide, transmembrane and cytoplasmic tail domains are all encoded by exon 5 in ZAA. In ZBA, this region in encoded by exons 5 and 6 where the trans- membrane region has been lost. In ZCA, the 4th exon is larger than for any other class I gene and encodes the a3 domain in addition to the connecting peptide, trans- membrane and cytoplasmic region. However this exon- intron organization is truly unorthodox. A sequence comparison of the ZBA sequence with the genomic region downstream of the ZCA a3 domain showed that a similar exon-intron organisation as in ZBA could be identified for ZCA as well. Thus the ESTs identified for the ZCA gene could be splicoforms. Phylogenetic analysis of fish MHC class I molecules The MHC class I amino acid sequences from the U-, S-, Z- and L-lineages in addition to MHC class I sequences from various fishes, man and shark were used to draw a phylogenetic tree (Figure 3 and Additional File 2). With bootstrap values ranging from 90-100% the U, Z, L and S sequences form distinct clusters. Most published fish class I sequences belong to the U-lineage, while the Z-lineage so far contains sequences from fewer teleost species including the pufferfish family, zebrafishes, carp and Japanese flounder. L-lineage class I molecules are so far only identified in zebrafish, fathead minnow and salmonids, while class I sequences from the S-lineage are only found in catfish and salmonids. Among the Atlantic salmon U-lineage class I sequences there is a close relationship between the Sasa-UBA and Sasa-ULA suggesting that ULA is a recent duplication of UBA. An ULA homolog is so far not identified in rainbow trout. Sasa-UDA and Sasa-UGA cluster on