1. From the Ensembl homepage, choose Human from the drop-down list and search for BRAF. Click through to the Gene tab view. Click on Orthologues at the left side of the page to see all the orthologous genes.

   There are 1:1 orthologues in 22 primates reported in the summary table.

   Search for Tarsier in the table below.

   The percentage of identical amino acids in the tarsier protein (the orthologue) compared with the gene of interest. i.e. human BRAF (the target species/gene) is 95.39%. This is known as the Target%id. The identity of the gene of interest (human BRAF) when compared with the orthologue (tarsier BRAF, the query species/gene) is 94.65% (the Query %id). Note the difference in the values of the Target and Query %id reflects the different protein lengths for the human and tarsier BRAF genes.

   Click on the View Sequence Alignments link in the Orthologue column to View Protein Alignment in Clustal W format.

   Conserved amino acids are indicated by asteriks. The alignment around the N-terminus looks poorer, when compared to the C-terminus end.

2. Click on Gene tree in the left hand menu. All of the primates are enclosed in a lilac box. Click on the furthest left node in the box to get a pop-up labelled Primates. Alternatively, scroll to the bottom of the page, and select Order from Collapse all the nodes at the taxonomic rank. Primates will appear as a red triangle. Click on Wasabi viewer in the pop-up menu to see the alignment. Scroll to position 450.

   Greater bamboo lemur, mouse lemur, Sumatran orangutan, crab-eating macaque, olive baboon, Bolivian squirrel monkey, white-tufted-ear marmoset and Ma’s night monkey all match the human sequence.

(a) Search for cow ABCC11 from the home page. Click on ABCC11 genomic coordinates 18:16590653-16667410:-1 in the search results to open the Location tab. Click on Alignments (image) at the left, and select the 43 eutherian mammals EPO multiple alignment by clicking on Select an alignment blue button. Scroll down to see the hidden and missing species.

All but 13 of the 43 mammals have an alignment at this region. ABCC11 gene model for the closely related Pecora species (cows, yak, goat, sheep and Yarkand deer) is longer when compared to the other species, with many additional exons at its 3’ end (left side of the image), which are absent in other taxa.

(b) Click on Alignments (text) in the left hand menu. The 43 eutherian mammals EPO alignment should be pre-selected. Scroll down to the table of alignment blocks. Sort the table by clicking on small arrows in the Location on Cow column header. The alignment blocks are now sorted by the genomic coordinates, with smalles coordiantes corresponding to the 3’ most end of ABCC11 (located on the reverse strand). Click on Block 3 to view the alignment.

Only 5 species have an alignment in this region, including cows, yak, goat, sheep and Yarkand deer, which is in agreement with our previous observation. Scroll up and click on Download alignment blue button, change File format to CLUSTALW, then Download.

(c) Click on Region in detail in the left hand menu. Turn on the 90 eutherian mammals EPO-Extended, Constrained elements and Conservation score for 90 eutherian mammals EPO-Extended tracks, all under the Comparative genomics in the Configure this page menu.

The 90 eutherian mammals EPO-Extended multiple alignment track is shown as pink block indicating that the whole region can be aligned at this locus. The GERP elements and GERP scores tracks show where the conserved sequence is located in the alignment. Conserved elements shown as pink boxes match up with exonic regions of the 5’-half of this cow gene (right side of the image). In general, exons tend to be highly conserved across taxa. Click on the track name and the i icon (information button) to read more about constrained elements (or any other data track).

1. Search for human RHO from the home page. Click on RHO genomic coordinates 3:129528639-129535344:1 in the search results to directly open the Location tab. Click Synteny at the left and change the species to Duck next to the image.

   Yes, there are multiple syntenic regions in duck to human chromosome 3, which is in the centre of this view. Duck chromosomes 1, 2, 7, 9, 13, and Z have syntenic regions to human chromosome 3.

2. Scroll down to the bottom of the page to see a list of homologous genes on both genomes.

   Human RHO is homologous to RHO (ENSAPLG00000005189) in duck. Click 15 upstream genes or 15 downstream genes to view neighbouring genes in this syntenic block. There are many neighbouring genes with homologues in duck. Human genes in this region are homologous to duck genes on chromosome 13, which is also indicated by the red boxes in the image above as this genomic block on human chromosome 3 is syntenic to duck chromosome 13.

(a) Select Human from the species selector drop-down list and type brca2 in the search box. Click Go. Click on 13:32315086-32400268:1 below BRCA2 (Human Gene) to go to the Region in detail page.

Click Configure this page in the side menu, then BLASTz/LASTz alignments under the Comparative genomics menu. Select Chicken, Chimpanzee, Mouse and Platypus in Normal style.

Yes, the degree of conservation does reflect the evolutionary relationship between human and the other species; the highest degree of conservation is found in chimp, followed by mouse, platypus and chicken, respectively.

Especially the exonic sequences of BRCA2 seem to be highly conserved between the various species, which is what is to be expected because these are supposed to be under higher selection pressure than intronic and intergenic sequences.

(b) Click Configure this page in the side menu, then Conservation regions under the Comparative genomics menu.

Select Conservation score and Constrained elements for 90 eutherian mammals EPO-Extended and 65 amniota vertebrates Mercator-Pecan.

Both the Conservation score and Constrained elements tracks largely correspond with the data seen in the pairwise alignment tracks; all exons of the BRCA2 gene show a high degree of conservation (note the UTRs which are not conserved).

(c) Click on exons of the golden transcript (ENST00000380152) to reveal their rank in transcript. Exon 15 can be found in the middle. Click on a constrained element in 65 way GERP elements track matching up with this exon.

Click on View alignments (text) in the pop-up menu, then Configure this page in the side menu. Select Show conservation regions to highlight bases matching in majority of the species in this alignment.

Exons are indicated by red lettering. All but Naja naja (Indian cobra) and Pseudonaja textilis (Eastern brown snake) have exonic sequence in this region.