Exercise solutions

Exercise 1 – Orthologues, paralogues and gene trees for the human BRAF gene

(a) Go to www.ensembl.org, choose human and search for BRAF. Click through to the Gene tab view.

On the gene tab, click on Orthologues at the left side of the page to see all the orthologous genes.

There are orthologues in 10 of the 11 primates represented in Ensembl. Only Vervet-AGM has no predicted BRAF orthologue.

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 69%. 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 62% (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.


(b) There is more than one way to get to the answer.

Option 1: Go to the Orthologues page and click on the marmoset orthologue to open the gene tab.

Click Genomic alignments at the left. Then select Select an Alignment: Human (Homo_sapiens) and click Apply. Choose Block 1 to get the largest block of aligned sequence, then click Display full alignment.

The red sequence is present in exons, so there is a gene in both species in this region. You can find where the start and stop codons are located if you configure this page and select START/STOP codons.

Option 2: Go to location tab of the marmoset BRAF gene and then click on  Region Comparison view at the left. Click on Select species or regions at the left and select Human (Homo_sapiens) then click Apply. You should see an alignment between the human BRAF gene region and the BRAF gene region for the marmoset.

(Note: To see a blue line connecting homologous genes in the Region Comparison view page, click on configure this page and under Comparative features select join genes. Zoom out on the location view to see blue lines connecting all the homologous genes between marmoset and human genes in that region).


Exercise 2 – Zebrafish orthologues

(a) Start in the  Location tab (region in detail) for sardh. Click on  Alignments (Image) at the left, and click Select an alignment to select the Multiple: 11 fish EPO_LOW_COVERAGE alignment in the pop-up view.

The zebrafish, cave fish, cod, tilapia, Amazon molly, platyfish, spotted gar, stickleback, medaka, fugu, and tetradon are shown in this region. All the species show a gene in the aligned region. This can also be seen in the  Alignments (text) page (the exons are highlighted in red).


(b) You can export the alignments from either the  Alignments (text) or  Alignments (image) pages in the  Location tab. Click on the blue  Download alignments button at the top of the text page, or the icon at the top of the image, and choose  ClustalW from the list.


(c) Click on  Region in detail in the left hand menu. Turn on the  multiple alignment,  constrained elements and  conservation score for  11 fish EPO tracks, all under the  Comparative genomics menu by  configuring the page.

The 11 fish EPO track just shows that the whole region for the sardh gene can be aligned among those eleven species of fish. The Constrained elements and Conservation score tracks show the conserved sequence is located where in the alignment.

Higher conservation regions match up with exonic regions (exons tend to be highly conserved) of the gene.

Hover over the the Track name and the Macintosh HD:Users:setup:Desktop:Online materials:help icon.png (information button) to read more about constrained elements (or any other data track).


Exercise 3 – Synteny

(a) Change the species to dog next to the image.

Yes, there are multiple syntenic regions in dog to human chromosome 3, which is in the centre of this view. Dog chromosomes 6, 20, 23, 31, 33, and 34 have syntenic regions to human chromosome 3.

(b) Scroll down to the bottom of the page.

There is a homologue in dog of human RHO. Click Centre on gene RHO to compare the genes between human and dog in this syntenic block.


Exercise 4 – Whole genome alignments

(a) Go to the Ensembl homepage (http://www.ensembl.org/).

Select Search: Human and type brca2 in the search box.

Click Go.

Click on 13:32315474-32400266:1 below BRCA2 (Human Gene).

You may want to turn off all tracks that you added to the display in the previous exercises as follows:

Click Configure this page in the side menu.

Click Reset configuration.

SAVE and close.


(b) Click Configure this page in the side menu

Click on BLASTZ/LASTZ alignments under the Comparative genomics menu. Select Chicken (Gallus gallus) - LASTZ_NET – NormalChimpanzee (Pan troglodytes) – LASTZ_NET – NormalMouse (Mus musculus) – LASTZ_NET – Normal and Platypus (Ornithorhynchus anatinus) - LASTZ_NET – Normal.

SAVE and close.

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.


(c) Click Configure this page in the side menu.

Click on Conservation regions under the Comparative genomics menu.

Select Conservation score for 53 eutherian mammals EPO_LOW_COVERAGEConservation score for 27 amniota vertebrates Pecan and Constrained elements for 53 amniota vertebrates Pecan.

SAVE and close.

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).


(d) Click on a constrained element (brown block).

Click on View alignments (text) in the pop-up menu.

Click Configure this page in the side menu.

Select Show conservation regions.

SAVE and close.

The conserved regions will be shown in light blue.



Bonus Exercise 5 – Exporting paralogues with BioMart

Start at ensembl.org/biomart/martview. Choose the ENSEMBL Genes database. Choose the Homo sapiens Genes dataset.

Click on Filters in the left panel. Under REGION: select Chromosome – 14. Under MULTI SPECIES COMPARISONS: select Homologue filters – Paralogous Human Genes: Only.

Click on Attributes in the left panel. Select Homologues from the six options at the top. Under GENE: deselect Ensembl Transcript ID and select Associated Gene Name. Under PARALOGUES: select Human Paralogue Ensembl Gene ID, Human Paralogue associated gene name, Last common ancestor with Human, %id. target Human gene identical to query gene and %id. query gene identical to target Human gene.

Click the Results button on the toolbar. Select View All rows as HTML or export all results to a file.