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Omics Techniques and their Application to Genomic Medicine, Queen Mary University of London, 21 January 2022

Course Details

Lead Trainer
Michal Szpak
Event Date
  Queen Mary University of London: Virtual
A short introduction to Ensembl genes and variants as part of the MSc in Genomic Medicine.

Demos and exercises

Species and genome assemblies

The front page of Ensembl is found at ensembl.org. It contains lots of information and links to help you navigate Ensembl:

At the top left you can see the current release number and what has come out in this release. To access old releases, scroll to the bottom of the page and click on View in archive site.

Click on the links to go to the archives. Alternatively, you can jump quickly to the correct release by putting it into the URL, for example e98.ensembl.org jumps to release 98.

Click on View full list of all species.

Click on the common name of your species of interest to go to the species homepage. We’ll click on Human.

Here you can see links to example pages and to download flatfiles. To find out more about the genome assembly and genebuild, click on More information and statistics.

Here you’ll find a detailed description of how to the genome was produced and links to the original source. You will also see details of how the genes were annotated.

The current genome assembly for human is GRCh38. If you want to see the previous assembly, GRCh37, visit our dedicated site grch37.ensembl.org.

Let’s take a look at the Ensembl Genomes homepage at ensemblgenomes.org.

Click on the different taxa to see their homepages. Each one is colour-coded.

You can navigate most of the taxa in the same way as you would with Ensembl, but Ensembl Bacteria has a large number of genomes, so needs slightly different methods. Let’s look at it in more detail.

There’s no full species list for bacteria as it would be hard to navigate with the number of species. To find a species, start to type the species name into the species search box. A drop down list will appear with possible species.

For example, to find a sub-strain of Clostridioides difficile start typing in the species name. Due to the autocomplete, you’ll see useful results as soon as you get to clostridio.

The drop down contains various strains of Clostridioides difficile. Let’s choose Clostridioides difficile 630. This will take us to another species homepage, where we can explore various features.

Unlike the human homepage, there is no prose description of the genome or gene annotation, as these pages were generated automatically.

Our newest genomes, such as those coming from the Darwin Tree of Life, are available rapid.ensembl.org with limited annotation.

Panda species data

(a) Go to the species homepage for Giant panda. What is the name of the genome assembly for Panda?

(b) Click on More information and statistics. How long is the Panda genome (in bp)? How many genes have been annotated?

(a) Select Giant panda from the drop down species list, or click on View full list of all Ensembl species, then choose Giant panda from the list.

The assembly is ASM200744v2 or GCA_002007445.2.

(b) Click on More information and statistics. Statistics are shown in the tables on the left.

The length of the genome is 2,444,060,653 bp.
There are 20,857 coding genes.

Available zebrafish assemblies

What previous assemblies are available for zebrafish?

Click on Zebrafish on the front page of Ensembl to go to the species homepage. Under Other assemblies three previous assembly names and the releases you can find them in are listed.

Assembly GRCz10 is available in the archived release 91, Zv9 in 79 and Zv8 in 54.

Mosquito species

(a) Go to Ensembl Metazoa. How many species of the genus Anopheles are there in Ensembl Metazoa?

(b) When was the current Anopheles gambiae genome assembly last revised?

(a) Go to metazoa.ensembl.org. Open the drop down list or click on View full list of all Ensembl Metazoa species. In a latin binomial species name, the first word represents the genus. Type Anopheles into the filter box in the top left to find all genomes with this word in the binomial.

There are 22 Anopheles species.

(b) Click on Anopheles gambiae, then on More information and statistics.

The genome was revised in February 2006.

Finding genomes with species search on Ensembl Bacteria

Mycobacterium tuberculosis H37Ra str. ATCC25177 is a clinical strain.

Go to Ensembl Bacteria and find the species Mycobacterium tuberculosis H37Ra str. ATCC25177 (Hint: type H37Ra into the Search for a genome box). How many coding genes does it have?

Go to bacteria.ensembl.org and start to type the name H37Ra into the search species box. It will autocomplete, allowing you to select Mycobacterium tuberculosis H37Ra str. ATCC25177 from the drop-down list. Click on More information and statistics.

Mycobacterium tuberculosis H37Ra str. ATCC25177 has 4034 coding genes and 48 non-coding.

Exploring genomic regions

Start at the Ensembl front page, ensembl.org. You can search for a region by typing it into a search box, but you have to specify the species.

To bypass the text search, you need to input your region coordinates in the correct format, which is chromosome, colon, start coordinate, dash, end coordinate, with no spaces for example: human 4:122868000-122946000. Type (or copy and paste) these coordinates into either search box.


Press Enter or click Go to jump directly to the Region in detail Page.

Click on the button to view page-specific help. The help pages provide text, labelled images and, in some cases, help videos to describe what you can see on the page and how to interact with it.

The Region in detail page is made up of three images, let’s look at each one in detail.

  1. The first image shows the chromosome:

The region we’re looking at is highlighted on the chromosome. You can jump to a different region by dragging out a box in this image. Drag out a box on the chromosome, a pop-up menu will appear.

If you wanted to move to the region, you could click on Jump to region (### bp). If you wanted to highlight it, click on Mark region (###bp). For now, we’ll close the pop-up by clicking on the X on the corner.

  1. The second image shows a 1Mb region around our selected region. This is always 1Mb in human, but the fixed size of this view varies between species. This view allows you to scroll back and forth along the chromosome.

You can also drag out and jump to or mark a region.

Click on the X to close the pop-up menu.

Click on the Drag/Select button to change the action of your mouse click. Now you can scroll along the chromosome by clicking and dragging within the image. As you do this you’ll see the image below grey out and two blue buttons appear. Clicking on Update this image would jump the lower image to the region central to the scrollable image. We want to go back to where we started, so we’ll click on Reset scrollable image.

  1. The third image is a detailed, configurable view of the region.

Here you can see various tracks, which is what we call a data type that you can plot against the genome. Some tracks, such as the transcripts, can be on the forward or reverse strand. Forward stranded features are shown above the blue contig track that runs across the middle of the image, with reverse stranded features below the contig. Other tracks, such as variants, regulatory features or conserved regions, refer to both strands of the genome, and these are shown by default at the very top or very bottom of the view.

You can use click and drag to either navigate around the region or highlight regions of interest, Click on the Drag/Select option at the top or bottom right to switch mouse action. On Drag, you can click and drag left or right to move along the genome, the page will reload when you drop the mouse button. On Select you can drag out a box to highlight or zoom in on a region of interest.

With the tool set to Select, drag out a box around an exon and choose Mark region.

The highlight will remain in place if you zoom in and out or move around the region. This allows you to keep track of regions or features of interest.

We can edit what we see on this page by clicking on the blue Configure this page menu at the left.

This will open a menu that allows you to change the image.

There are thousands of possible tracks that you can add. When you launch the view, you will see all the tracks that are currently turned on with their names on the left and an info icon on the right, which you can click on to expand the description of the track. Turn them on or off, or change the track style by clicking on the box next to the name. More details about the different track styles are in this FAQ: http://www.ensembl.org/Help/Faq?id=335.

You can find more tracks to add by either exploring the categories on the left, or using the Find a track option at the top left. Type in a word or phrase to find tracks with it in the track name or description.

Let’s add some tracks to this image. Add:

  • Proteins (mammal) from UniProt – Labels
  • 1000 Genomes - All - short variants (SNPs and indels) – Normal

Now click on the tick in the top left hand to save and close the menu. Alternatively, click anywhere outside of the menu. We can now see the tracks in the image. The proteins track is stranded, so you will see two tracks, one above and one below the contig, representing the proteins mapped to the forward and reverse strands respectively. The variants track is not stranded, so is found near the bottom of the image.

If the track is not giving you can information you need, you can easily change the way the tracks appear by hovering over the track name then the cog wheel to open a menu. To make it easier to compare information between tracks, such as spotting overlaps, you can move tracks around by clicking and dragging on the bar to the left of the track name.

Now that you’ve got the view how you want it, you might like to show something you’ve found to a colleague or collaborator. Click on the Share this page button to generate a link. Email the link to someone else, so that they can see the same view as you, including all the tracks you’ve added. These links contain the Ensembl release number, so if a new release or even assembly comes out, your link will just take you to the archive site for the release it was made on.

To return this to the default view, go to Configure this page and select Reset configuration at the bottom of the menu.

Exploring a genomic region in human

(a) Go to the region from 31,937,000 to 32,633,000 bp on human chromosome 13. On which cytogenetic band is this region located? How many contigs make up this portion of the assembly (contigs are contiguous stretches of DNA sequence that have been assembled solely based on direct sequencing information)?

(b) Zoom in on the BRCA2 gene.

(c) Configure this page to turn on the LTR (repeat) track in this view. What tool was used to annotate the LTRs according to the track information? How many LTRs can you see within the BRCA2 gene? Do any overlap exons?

(d) Create a Share link for this display. Email it to your neighbour. Open the link they sent you and compare. If there are differences, can you work out why?

(e) Export the genomic sequence of the region you are looking at in FASTA format.

(f) Turn off all tracks you added to the Region in detail page.

(a) Go to the Ensembl homepage.

Select Search: Human and type 13:31937000-32633000 in the text box (or alternatively leave the Search drop-down list like it is and type human 13:31937000-32633000 in the text box). Click Go.

This genomic region is located on cytogenetic band q13.1. It is made up of eight contigs, indicated by the alternating light and dark blue coloured bars in the Contigs track. Note that KF455761.1 is a tiny contig that splits AL137143.8 in two.

(b) Draw with your mouse a box encompassing the BRCA2 transcripts. Click on Jump to region in the pop-up menu.

(c) Click Configure this page in the side menu (or on the cog wheel icon in the top left hand side of the bottom image).

Go into Repeats in the left-hand menu then select LTR. Click on the (i) button to find out more

Repeat Masker was used to annotate LTRs onto the genome.

Save and close the new configuration by clicking on ✓ (or anywhere outside the pop-up window).

There are seven LTRs overlapping BRCA2, none of them overlap exons.

(d) Click Share this page in the side menu. Select the link and copy. Get your neighbour’s email address and compose an email to them, paste the link in and send the message.

When you receive the link from them, open the email and click on your link. You should be able to view the page with the new configuration and data tracks they have added to in the Location tab. You might see differences where they specified a slightly different region to you, or where they have added different tracks.

Here is the Share link from the video answer: http://may2021.archive.ensembl.org/Homo_sapiens/Share/71a173bba78f0dbe03e48d3240424943?redirect=no;mobileredirect=no

(e) Click Export data in the side menu. Leave the default parameters as they are (FASTA sequence should already be selected). Click Next>. Click on Text.

Note that the sequence has a header that provides information about the genome assembly (GRCh38), the chromosome, the start and end coordinates and the strand. For example:

>13 dna:chromosome chromosome:GRCh38:13:32311910:32405865:1

(f) Click Configure this page in the side menu. Click Reset configuration. Click ✓.

Exploring CRISPR sites in a human genomic region

You want to do some CRISPR manipulation of the human SMC3 gene. You’re looking for a CRISPR editing site within the locus 10:110578600-110578700.

(a) Go to the locus above and turn on the CRISPR track. How many CRISPR sites can you see in this locus?

(b) Do any of the CRISPR sites overlap any phenotype causing variants? What are the identifiers of these sites and variants?

(c) Mark the region of the negative strand CRISPR site that overlaps these variants, then zoom out to see the whole SMC3 gene. What exon number is your CRISPR site found in the SMC3-201 transcript?

(a) Go to the Ensembl homepage.

Select Search: Human and type 10:110578600-110578700 in the text box (or alternatively leave the Search drop-down list like it is and type human 10:110578600-110578700 in the text box). Click Go.

Click Configure this page. Type crispr in the Find a track text box. Select CRISPR Cas9 in Labels.

There are five positive strand and three negative strand CRISPR sites.

(b) Click on the variants and CRISPR sites to get their identifiers.

Forward stranded CRISPR site 1074131233 overlaps rs78663177. 1074131234, 1074131235 and 1074131236 overlap bunch of phenotype associated variants including rs113411202, rs1064797151, rs779773957, rs1590553017, rs972620847 and rs748876063. 1074131234 also overlaps rs1554882313.

(c) Click and drag a box around the site, then select Mark region. In the overview above, click and drag a box around the SMC3 gene then select Jump to region. Count the exons to get the number where the marked region is found.

The site is found in exon 7.

Exploring assembly exceptions in human

(a) Go to the region 21:32630000-32870000 in human. What is the red highlighted region? What is its name?

(b) Can you see the assembly exceptions in the chromosome overview at the top? How many regions with assembly exceptions are there on chromosome 21?

(c) Can you compare this assembly exception with the reference? What is different between this assembly exception and the version on the primary assembly?

(a) Go to the Ensembl homepage.

Select Search: Human and type 21:32630000-32870000 in the text box (or alternatively leave the Search drop-down list like it is and type human 21:32630000-32870000 in the text box). Click Go.

You will see a red highlighted region in the middle of this region. Click on the thin dark red bar in any of the three views to see the label _CHR_HSCHR21_3_CTG1_1:32769079-32843731__. Click on _What are assembly exceptions? to open a new window which explains assembly exceptions.

(b) Assembly exceptions are marked in the chromosome view at the top.

There are seven haplotypes on chromosome 21 and one patch.

(c) Another option in the drop-down is Compare with reference. Click on this.

Scroll down the page to see the comparison between the haplotype and primary assembly. Aligned sequences are highlighted in pink and linked together in green.

The assembly exception CHR_HSCHR21_3_CTG1_1 contains an extra region compared to the primary assembly.

Exploring a genomic region in rice

(a) Go to the region 1:405000-453000 in Oryza sativa Japonica.

(b) Turn on the AGILENT:G2519F-015241 microarray track. Are there any oligo probes that map to this region?

(c) Highlight the region around any reverse strand probes you can see. Do they map to any transcripts?

(a) Go to the Ensembl Plants homepage.

Select Search: Oryza sativa Japonica and type 1:405000-453000. Click Go.

(b) Click on Configure this page to open the menu. You can find the AGILENT:G2519F-015241 track under Oligo probes in the left hand menu, or by using the Find a track box at the top right. Turn on the track then save and close the menu.

As the AGILENT:G2519F-015241 track is stranded, it appears at the top and bottom of the view, in green.

There are five probes mapped to this region on the positive strand and one probe on the reverse strand.

(c) Drag a box around the reverse strand probe then click on Mark region to highlight.

The highlighted region maps to two transcripts: Os01t0107900-02 and Os01t0107900-01

Exploring a genomic region in Salmonella enterica

(a) Search for the Salmonella enterica subsp. enterica serovar Typhi str. Ty2 (Hint: type Ty2 into the Search for a genome box).

(b) Go to the region Chromosome:2000605-2009742.

(c) How many genes are annotated in this region? How many are on the forward strand? How many are on the reverse strand?

(a) Go to the Ensembl Bacteria homepage.

Type Ty2 into the Search for a Genome box. Click on the auto-completed genome name to navigate to the species homepage.

(b) Type Chromosome:2000605-2009742 into the search box. Click Go.


There are eight genes annotated in this region, all on the reverse strand.

Genes and transcripts

You can find out lots of information about Ensembl genes and transcripts using the browser. If you’re already looking at a region view, you can click on any transcript and a pop-up menu will appear, allowing you to jump directly to that gene or transcript.

Alternatively, you can find a gene by searching for it. You can search for gene names or identifiers, and also phenotypes or functions that might be associated with the genes.

We’re going to look at the human UQCRQ gene. From ensembl.org, type UQCRQ into the search bar and click the Go button. You will get a list of hits with the human gene at the top.

Where you search for something without specifying the species, or where the ID is not restricted to a single species, the most popular species will appear first, in this case, human, mouse and zebrafish appear first. You can restrict your query to species or features of interest using the options on the left.

The gene tab

Click on the gene name or Ensembl ID. The Gene tab should open:

This page summarises the gene, including its location, name and equivalents in other databases. At the bottom of the page, a graphic shows a region view with the transcripts. We can see exons shown as blocks with introns as lines linking them together. Coding exons are filled, whereas non-coding exons are empty. We can also see the overlapping and neighbouring genes and other genomic features.

There are different tabs for different types of features, such as genes, transcripts or variants. These appear side-by-side across the blue bar, allowing you to jump back and forth between features of interest. Each tab has its own navigation column down the left hand side of the page, listing all the things you can see for this feature.

Let’s walk through this menu for the gene tab. How can we view the genomic sequence? Click Sequence at the left of the page.

The sequence is shown in FASTA format. The FASTA header contains the genome assembly, chromosome, coordinates and strand (1 or -1) – this gene is on the positive strand.

Exons are highlighted within the genomic sequence, both exons of our gene of interest and any neighbouring or overlapping gene. By default, 600 bases are shown up and downstream of the gene. We can make changes to how this sequence appears with the blue Configure this page button found at the left. This allows us to change the flanking regions, add variants, add line numbering and more. Click on it now.

Once you have selected changes (in this example, Show variants, 1000 Genomes variants and Line numbering) click at the top right.

You can download this sequence by clicking in the Download sequence button above the sequence:

This will open a dialogue box that allows you to pick between plain FASTA sequence, or sequence in RTF, which includes all the coloured annotations and can be opened in a word processor. If you want run a sequence analysis tool, download as FASTA sequence, whereas if you want to analyse the sequence visually, RTF is best for this. This button is available for all sequence views.

To find out what the protein does, have a look at GO terms from the Gene Ontology consortium. There are three pages of GO terms, representing the three divisions in GO: Biological process (what the protein does), Cellular component (where the protein is) and Molecular function (how it does it). Click on GO: Biological process to see an example of the GO pages.

Here you can see the functions that have been associated with the gene. There are three-letter codes that indicate how the association was made, as well as links to the specific transcript they are linked to.

We also have links out to other databases which have information about our genes and may focus on other topics that we don’t cover, like Gene Expression Atlas or OMIM. Go up the left-hand menu to External references:

Demo: The transcript tab

We’re now going to explore the different transcripts of UQCRQ. Click on Show transcript table at the top.

Here we can see a list of all the transcripts of UQCRQ with their identifiers, lengths, biotypes and flags to help you decide which ones to look at.

If we were to only choose one transcript to analyse, we would choose UQCRQ-203 because it is the MANE Select and Ensembl Canonical. This means it is both 100% identical to the RefSeq transcript NM_014402.5 and both Ensembl and NCBI agree that it is the most biologically important transcript.

Click on the ID, ENST00000378670.8.

You are now in the Transcript tab for UQCRQ-203. We can still see the gene tab so we can easily jump back. The left hand navigation column provides several options for the transcript UQCRQ-203 - many of these are similar to the options you see in the gene tab, but not all of them. If you can’t find the thing you’re looking for, often the solution is to switch tabs.

Click on the Exons link. This page is useful for designing RT-PCR primers because you can see the sequences of the different exons and their lengths.

You may want to change the display (for example, to show more flanking sequence, or to show full introns). In order to do so click on Configure this page and change the display options accordingly.

Now click on the cDNA link to see the spliced transcript sequence with the amino acid sequence. This page is useful for mapping between the RNA and protein sequences, particularly genetic variants.

UnTranslated Regions (UTRs) are highlighted in dark yellow, codons are highlighted in light yellow, and exon sequence is shown in black or blue letters to show exon divides. Sequence variants are represented by highlighted nucleotides and clickable IUPAC codes are above the sequence.

Next, follow the General identifiers link at the left. Just like the External References page in the gene tab, this page shows links out to other databases such as RefSeq, UniProtKB, PDBe and others, this time linked to the transcript or protein product, rather than the gene.

If you’re interested in protein domains, you could click on Protein summary to view domains from Pfam, PROSITE, Superfamily, InterPro, and more. These are all plotted against the transcript sequence, with the exons shown in alternating shades of purple at the top of the page. Alternatively, you can go to Domains & features to see a table of the same information.

You can also see the structure of the protein from the PDB by clicking on PDB 3D Protein model.

This uses LiteMol to show a 3D protein. You can use all the normal controls that you would use with LiteMol, plus plot Ensembl features like Exons and variants onto the structure using the options on the right. We allow you to see the top ten PDB models for this protein, based on coverage and quality scores, you can choose which at the top of the viewer.

Exploring the human MYH9 gene

(a) Find the human MYH9 (myosin, heavy chain 9, non-muscle) gene, and go to the Gene tab.

  • On which chromosome and which strand of the genome is this gene located?
  • How many transcripts (splice variants) are there and how many are protein coding?
  • What is the longest transcript, and how long is the protein it encodes?
  • Which transcript would you take forward for further study?

(b) Click on Phenotypes at the left side of the page. Are there any diseases associated with this gene, according to MIM (Mendelian Inheritance in Man)?

(c) What are some functions of MYH9 according to the Gene Ontology consortium? Have a look at the GO pages for this gene.

(d) In the transcript table, click on the transcript ID for MYH9-201, and go to the Transcript tab.

  • How many exons does it have?
  • Are any of the exons completely or partially untranslated?
  • Is there an associated sequence in UniProtKB/Swiss-Prot? Have a look at the General identifiers for this transcript.

(e) Are there microarray (oligo) probes that can be used to monitor ENST00000216181 expression?

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

Select Search: Human and type MYH9. Click Go.

Click on either the Ensembl ID ENSG00000100345 or the HGNC official gene name MYH9.

  • Chromosome 22 on the reverse strand.
  • Ensembl has 23 transcripts annotated for this gene, of which six are protein coding.
  • The longest transcript is MYH9-215 and it codes for a protein of 1,981 amino acids
  • MYH9-201 is the transcript I would take forward for further study, as it is the MANE Select.

(b) Click on Phenotypes at the left to see the associated phenotypes. There is a large table of phenotypes. To see only the ones from MIM, type mim into the filter box at the top right of the table.

These are some of the phenotypes associated with MYH9 according to MIM: autosomal dominant deafness and Macrothrombocytopenia and granulocyte inclusions with or without nephritis or sensorineural hearing loss. Click on the records for more information.

(c) > The Gene Ontology project (http://www.geneontology.org/) maps terms to a protein in three classes: biological process, cellular component, and molecular function. Meiotic spindle organisation, cell morphogenesis, and cytokinesis are some of the roles associated with MYH9.

(d) Click on ENST00000216181.11

  • It has 41 exons, shown in the Transcript summary.

Click on the Exons link in this side menu.

  • Exon 1 is completely untranslated, and exons 2 and 41 are partially untranslated (UTR sequence is shown in orange). You can also see this in the cDNA view if you click on the cDNA link in the left side menu.
  • P35579-1 from UniProt/Swiss-Prot matches the translation of the Ensembl transcript. Click on P35579-1 to go to UniProtKB, or click align for the alignment.

(e) Click on Oligo probes in the side menu.

Probesets from Affymetrix, Agilent, Codelink, Illumina, and Phalanx match to this transcript sequence. Expression analysis with any of these probesets would reveal information about the transcript. Hint: this information can sometimes be found in the ArrayExpress Atlas: www.ebi.ac.uk/arrayexpress/

Finding a gene associated with a phenotype

Phenylketonuria is a genetic disorder caused by an inability to metabolise phenylalanine in any body tissue. This results in an accumulation of phenylalanine causing seizures and mental retardation.

(a) Search for phenylketonuria from the Ensembl homepage and narrow down your search to only genes. What gene is associated with this disorder?

(b) How many protein coding transcripts does this gene have? View all of these in the transcript comparison view.

(c) What is the MIM gene identifier for this gene?

(d) Go to the MANE Select transcript and look at its 3D structure. In the model 2pah, how many protein molecules can you see?

(a) Start at the Ensembl homepage (http://www.ensembl.org).

Type phenylketonuria into the search box then click Go. Choose Gene from the left hand menu.

The gene associated with this disorder is PAH, phenylalanine hydroxylase, ENSG00000171759.

(b) If the transcript table is hidden, click on Show transcript table to see it.

There are six protein coding transcripts.

Click on Transcript comparison in the left hand menu. Click on Select transcripts. Either select all the transcripts labelled protein coding one-by-one, or click on the drop down and select Protein coding. Close the menu.

(c) Click on External references.

The MIM gene ID is 612349.

(d) Open the transcript table and click on the ID for the MANE Select: ENST00000553106.6. Go to PDB 3D protein model in the left-hand menu.

The model 2pah is shown by default. It has two protein molecules in it. You may need to rotate the model to see this clearly.

Exploring the mouse Dpp6 gene

Genetic variation in the dipeptidylpeptidase 6 Gene (DPP6) in humans has previously been strongly associated with amyotrophic lateral sclerosis (ALS), a lethal disorder caused by progressive degeneration of motor neurons in the brain.

(a) Search for the Dpp6 gene in mouse and click on the ENSMUST00000071500 transcript to open the transcript tab. How many exons make up this transcript?

(b) Click on Exons to display the exon sequences of the transcript. Which exon contains the translation start? What is the exon ID of the largest exon? What is the start and end phase of exon 2?

(c) Click on Protein summary. How many protein domains or features fall within the second exon? What is the PFAM protein domain at the C-terminus of the protein and how many exons does it fall into? Which amino acid positions does the domain above cover?

(d) Click on Domains and features. Which domains are associated with Pfam? How many genes in the mouse genome have the IPR002469 domain? What chromosomes are these genes found on?

(a) Go to the Ensembl homepage.

Select Search: Mouse and type Dpp6. Click Go.

Click on either the Ensembl ID ENSMUSG00000061576 or the MGI official gene name Dpp6. From the transcript table, click on the link for transcript ENSMUST00000071500 to open the transcript tab.

ENSMUST00000071500 consists of 26 exons.

(b) Click on Exons, which can be found on the left of the page. The translation start is found in the first exon (ENSMUSE00000725552), shown in dark blue text.

The largest exon is the final exon (856 bp), which has the exon ID ENSMUSE00000773588.

Exon 2 has a start and end phase of 0 and 1 respectively, which means that the codon at the start of the exon starts at the first nucleotide and the codon at the end of the exon ends at nucleotide 2. Notice that the end phase of each exon is the same as the start phase of the next exon.

(c) Click on Protein Summary in the menu on the left hand side of the page. Alternating exons are shown on the protein as different shades of purple.

There are two predicted protein domains that fall within the second exon: low complexity (seg) and a transmembrane helix.

Click on a domain or feature to view further information.

The C-terminal PFAM domain is Peptidase_S9 (PF00326), which spans or partially spans seven exons, covering amino acid positions 582-787.

(d) Click on Domains & features.

Looking at the Domains table you should notice that there are two domains associated with Pfam: PF00326 and PF00930.

Click on Display all genes with this domain next to IPR002469. This should now display the genes that have the IPR002469 domain located on the karyotype and as a table.

Six genes have this domain and they are found on chromosomes 1, 2, 5, 9 and 17.

Exploring a plant gene (Vitis vinifera, grape)

Start in http://plants.ensembl.org/index.html and select the Vitis vinifera genome.

(a) What GO: biological process terms are associated with the MADS4 gene?

(b) Go to the transcript tab for the only transcript, Vv01s0010g03900.t01. How many exons does it have? Which one is the longest? How much of that is coding?

(c) What domains can be found in the protein product of this transcript? How many different domain prediction methods agree with each of these domains?

(a) Go to http://plants.ensembl.org/index.html.

Select Vitis vinifera from the drop down menu All genomes – select a species or click on View full list of all Ensembl Plants species and then choose V. vinifera.

Type MADS4 and click on the gene link VIT_01s0010g03900. Click on GO: Biological process in the side menu.

There are seven terms listed including GO:0006351, transcription, DNA-templated, and GO:0006355, regulation of transcription, DNA-templated.

(b) Click on the transcript named Vv01s0010g03900.t01 (or on the Transcript tab). Click on Exons in the left hand menu.

There are eight exons. Exon 8 is longest with 303 bp, of which 13 are coding.

(c) Click on either Protein Summary or Domains & features in the left hand menu to see graphically or as a table respectively.

A MADS-box domain near the N-terminus is identified by eight domain prediction methods. A K-box domain near the C-terminus is identified by two. Two coiled-coils are identified by one.


In any of the sequence views shown in the Gene and Transcript tabs, you can view variants on the sequence. You can do this by clicking on Configure this page from any of these views.

Let’s take a look at the Gene sequence view for HBB in human. Search for HBB and go to the Sequence view.

If you can’t see variants marked on this view, click on Configure this page and select Show variants: Yes and show links. You may also wish to add a filter to the variants to allow them to load more quickly, we’ll add Filter variants by evidence status: 1000Genomes.

Find out more about a variant by clicking on it.

You can add variants to all other sequence views in the same way.

You can go to the Variation tab by clicking on the variant ID. For now, we’ll explore more ways of finding variants.

To view all the sequence variants in table form, click the Variant table link at the left of the gene tab.

You can filter the table to only show the variants you’re interested in. For example, click on Consequences: All, then select the variant consequences you’re interested in. For display purposes, the table above has already been filtered to only show missense variants.

You can also filter by the different pathogenicity scores and MAF, or click on Filter other columns for filtering by other columns such as Evidence or Class.

The table contains lots of information about the variants. You can click on the IDs here to go to the Variation tab too.

You can also see the phenotypes associated with a gene. Click on Phenotype in the left hand menu.

Open the transcript table and go to HBB-201 ENST00000335295, then click on Haplotypes in the left hand menu.

The Haplotypes view in the transcript tab shows you the actual protein and CDS sequences in 1000 Genomes individuals. This is possible because the 1000 Genomes study has phased genotypes, so we know which alleles occur on which of the chromosome pairs. The table lists all the versions of the protein that occur along with their frequencies, including the reference sequence and sequences with one or more alternative alleles.

Click on one of the haplotypes, we’ll go for 18K>*,​19del{130}, to find out more about it. Here you will see the frequency in the 1000 Genomes subpopulations, the sequence and the 1000 Genomes individuals where this protein is found.

Let’s have a look at variants in the Location tab. Click on the Location tab in the top bar.

Configure this page and open Variation from the left-hand menu.

There are various options for turning on variants. You can turn on variants by source, by frequency, presence of a phenotype or by individual genome they were isolated from. You can also turn on genotyping chips.

Let’s have a look at a specific variant. If we zoomed in we could see the variant rs334 in this region, however it’s easier to find if we put rs334 into the search box. Click through to open the Variation tab.

The icons show you what information is available for this variant. Click on Genes and regulation, or follow the link on the left.

This page illustrates the genes the variant falls within and the consequences on those genes, including pathogenicity predictors. It also shows data from GTEx on genes that have increased/decreased expression in individuals with this variant, in different tissues. Finally, regulatory features and motifs that the variant falls within are shown.

We can also see the variant in the protein structure by clicking on 3D Protein model.

This is a LiteMol viewer, where you can rotate and zoom in on the structure. The variant location is highlighted, so you can see where it lands within the structure.

Let’s look at population genetics. Click on Population genetics in the left-hand menu.

The population allele frequencies are shown by study, including 1000 Genomes and gnomAD. Where genotype frequencies are available, these are shown in the tables.

There are big differences in allele frequencies between populations. Let’s have a look at the phenotypes associated with this variant to see if they are known to be specific to certain human populations. Click on Phenotype Data in the left-hand menu.

This variant is associated with various phenotypes, including sickle cell and malaria resistance. These phenotype associations come from sources including the GWAS catalog, ClinVar, Orphanet and OMIM. Where available, there are links to the original paper that made the association, the allele that is associated with the phenotype and p-values and other statistics.

Human population genetics and phenotype data

The SNP rs1738074 in the 5’ UTR of the human TAGAP gene has been identified as a genetic risk factor for a few diseases.

(a) In which transcripts is this SNP found?

(b) What is the least frequent genotype for this SNP in the Yoruba (YRI) population from the 1000 Genomes phase 3?

(c) What is the ancestral allele? Is it conserved in the 90 eutherian mammals?

(d) With which diseases is this SNP associated? Are there any known risk (or associated) alleles?

(a) Please note there is more than one way to get this answer. Either go to the Variation Table for the human TAGAP gene, and Filter variants to the 5’UTR, or search Ensembl for rs1738074 directly.

Once you’re in the Variation tab, click on the Genes and regulation link or icon.

This SNP is found in four transcripts of TAGAP. It is also intronic to nine non-coding transcripts and up/downstream to 14 non-coding transcripts.

(b) Click on Population genetics at the left of the variation tab. (Or, click on Explore this variation at the left and click the Population genetics icon.)

In Yoruba (YRI), the least frequent genotype is CC at the frequency of 5.6%.

(c) Click on Phylogenetic context.

The ancestral allele is T and it’s inferred from the alignment in primates.

Select the 91 eutherian mammals EPO-Extended alignment and click on Apply.

A region containing the SNP (highlighted in red and placed in the centre) and its flanking sequence are displayed. The T allele is conserved in all but three of the eutherian mammals displayed.

(d) Click Phenotype Data at the left of the Variation page.

This variation is associated with multiple sclerosis, celiac and white blood cell count. There are known risk alleles for both multiple sclerosis and celiac and the corresponding P values are provided. The allele A is associated with celiac disease. Note that the alleles reported by Ensembl are T/C. Ensembl reports alleles on the forward strand. This suggests that A was reported on the reverse strand in the original paper. Similarly, one of the alleles reported for Multiple sclerosis is G.

Exploring a SNP in human

The missense variation rs1801133 in the human MTHFR gene has been linked to elevated levels of homocysteine, an amino acid whose plasma concentration seems to be associated with the risk of cardiovascular diseases, neural tube defects, and loss of cognitive function. This SNP is also referred to as ‘A222V’, ‘Ala222Val’ as well as other HGVS names.

(a) Find the page with information for rs1801133.

(b) Is rs1801133 a Missense variation in all transcripts of the MTHFR gene? What is the amino acid change?

(c) Why are the alleles for this variation in Ensembl given as G/A and not as C/T, as in the literature?

(d) What is the major allele of rs1801133 in different populations?

(e) In which paper(s) is the association between rs1801133 and homocysteine levels described?

(f) According to the data imported from dbSNP, the ancestral allele for rs1801133 is G. Ancestral alleles in dbSNP are based on a comparison between human and chimp. Does the sequence at this same position in other primates confirm that the ancestral allele is G?

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

Type rs1801133 in the Search box, then click Go. Click on rs1801133.

(b) Click on Genes and Regulation in the side menu (or the Genes and Regulation icon).

No, rs1801133 is Missense variant in nine MTHFR transcripts. Please note that this variant is multialleleic with two alternative alleles - as this table displays one consequence per row, each transcript is listed twice.

The amino acid change is A/V for allele A, and A/G for allele C.

(c) In Ensembl, the alleles of rs1801133 are given as G/A/C because these are the alleles in the forward strand of the genome. In the literature, the alleles are given as C/T/G because the MTHFR gene is located on the reverse strand. The alleles in the actual gene and transcript sequences are C/T/G. In Ensembl, the allele that is present in the reference genome assembly is always put first.

(d) Click on Population genetics in the side menu.

In all populations but one, the allele G is the major one. The exception is CLM (Colombian in Medellin; 1000 Genomes).

(e) Click on Phenotype Data in the left hand side menu.

The specific studies where the association was originally described is given in the Phenotype Data table. Links between rs1801133 and homocysteine levels were described in four papers. Click on the pubmed IDs PMID:20031578, PMID:23696881, PMID:30339177 and PMID:23824729 for more details.

(f) Click on Phylogenetic Context in the side menu.

Select Alignment: 9 primates EPO and click Apply.

Gorilla, bonobo, chimp, macaque, gibbon, vervet, crab-eating macaque and mouse lemur all have a G in this position.

Exploring a SNP in mouse

Madsen et al in the paper ‘Altered metabolic signature in pre-diabetic NOD mice’ (PloS One. 2012; 7(4): e35445) have described several regulatory and coding SNPs, some of them in genes residing within the previously defined insulin dependent diabetes (IDD) regions. The authors describe that one of the identified SNPs in the murine Xdh gene (rs29522348) would lead to an amino acid substitution and could be damaging as predicted as by SIFT (http://sift.jcvi.org/).

(a) Where is the SNP located (chromosome and coordinates)?

(b) What is the HGVS recommendation nomenclature for this SNP?

(c) Why does Ensembl put the C allele first (C/T)?

(d) Are there differences between the genotypes reported in NOD/LTJ and BALB/cByJ, according to the PERLGEN panel?

(a) Go to www.ensembl.org, type rs29522348 in the search box. Click on rs29522348 (Mouse Variation).

SNP rs29522348 is located on 17:74231988. In Ensembl, its alleles are provided as in the forward strand.

(b) Click on HGVS names to reveal information about HGVS nomenclature.

This SNP has got five HGVS names, one at the genomic DNA level (NC_000083.7:g.74231988C>T), three at the transcript level (ENSMUST00000024866.6:c.721G>A, ENSMUST00000233162.2:n.738G>A and ENSMUST00000233621.2:c.*284G>A) and one at the protein level (ENSMUSP00000024866.5:p.Val241Ile).

(c) In Ensembl, the allele that is present in the reference genome assembly is always put first (C is the allele for the reference mouse genome, strain C57BL/6J).

(d) Click on Sample genotypes is the left hand side menu. In the summary of genotypes by population, click on Show for PERLEGEN:MM_PANEL2, or search for the two strain names. There are indeed differences between the genotypes reported in those two different strains. The genotype reported in NOD/LTJ is T|T whereas in BALB/cByJ the genotype is C|C.

Variation data in the tomato (S. lycopersicum) genome

(a) Find the Solyc02g084570.3 gene in tomato and go to its Location tab. Can you see the variation track?

(b) Zoom in around the last exon of this gene. What are the different types of variants seen in that region? What are the locations of any splice region variants mapped in the region?

(a) Search for Solyc02g084570.3 and click on the Location link in the results page. The variation track is shown at the bottom of the region.

(b) Zoom in around the last exon of this gene by drawing a box in the respective region. Please note the gene is on the reverse strand, so the last exon will be on the left hand side of that image.

The variation legend is shown at the bottom of the page, telling you what the colours mean.

The types of variants seen in that region are 3’ UTR variants, missense variants, synonymous variants and splice region variants.

Splice region variants are shown in orange. Click on the variants to get additional information on that variant including location.

The variants are found at 2:48285642 and 2:48285640-48285641.

Exploring VNTR in human

Variable number tandem repeats (VNTRs) show high variation in the number of repeats in the population and are commonly used in forensics (DNA fingerprinting) and to study genetic diversity. (a) Go to the region from 3074666 to 3075100 bp on human chromosome 4. Which gene does it overlap? Which exon of this gene falls in this region?

(b) Configure this page to turn on Repeats (low), Simple repeats (Repeats (low)) and Tandem repeats (TRF) tracks in this view. Can you see any repeats in this exon? What tools were used to annotate the repeats according to the track information?

(c) Zoom in on the polyglutamine (PolyQ) tract or (CAG)n to see its sequence. How many CAG repeats can you see in the human reference assembly? Does this tract overlap any phenotype-associated variants? What is the identifier of this variant?

(d) Go to the variant tab of the phenotype-associated variant. What is the consequence ontology of this variant? Does the reference allele match the number of repeats you have just counted? What is the shortest and longest allele?

(e) Are there any phenotypes associated with this variant?

(a) Select Search: Human and type 4:3074666-3075100 in the text box (or alternatively type human 4:3074666-3075100 in the text box). Click Go.

Click on the golden transcript falling in this region. You can see it’s exon 1 of 67 of the huntingtin gene (HTT).

(b) Click Configure this page in the side menu then select: Repeats (low), Simple repeats (Repeats (low)) and Tandem repeats (TRF).

There are two types of tandem repeats in this exon: polyglutamine (PolyQ) tract or (CAG)n and polyproline (PolyP) tract or (CCG)n; annotated by two different methods. Click on the track names to find more about the tools used for annotation: RepeatMasker and Tandem Repeats Finder.

(c) Draw with your mouse a box around the polyglutamine (PolyQ) tract or (CAG)n. Click on Jump to region in the pop-up menu.

There are 19 CAG repeats in the human reference sequence overlapping rs71180116 indicated by a pink bar in the All phenotype-associated - short variants (SNPs and indels) track.

(d) Click on the rs71180116 ID to go to the variant tab. You can see in the summary page that this variant is classified as an inframe insertion. Either click + to show all of the alleles in the summary page or go to the Genes and regulation table. This variant has many alternative alleles which differ in the number of repeats. The first allele in the expanded Alleles section of the summary page or the first allele in the Codons column in the Genes and regulation table is the reference allele. It is composed of 19 CAG repeats just as in the Region in detail view. The shortest allele has 7 repeats, the longest has 55 repeats.

(e) Click on Phenotype data in the side menu. This variant is associated with Huntington disease, a trinucleotide repeat disorder (polyQ disease) caused by a pathogenic number of CAG repeats (above 36 copies) in a coding region of HTT.

Variation data in fungi

(a) How many species in Ensembl Fungi have variation data?

(b) Select Fusarium oxysporum and search for FOXG_13574T0 gene. One of its upstream variants is SNP tmp_10_6610. What are the possible alleles for this polymorphic position? Which one is on the reference genome?

(c) What is the most frequent allele at this position?

(d) Which samples have the genotypes C|T and T|T?

(a) Go to Ensembl Fungi, click on View full list of all species. Click on the upward triangle next to the Variation database column to sort the table by species with variation data.

The table shows that we have eight fungi species currently with variation databases.

(b) Click on Fusarium oxysporum in the table and on the species page search for FOXG_13574T0. From the Gene tab, click on Variant table and then scroll down to find tmp_10_6610 or use the table search box to find it.

The alleles are C/T, where C is the reference allele.

(c) Click on tmp_10_6610 in the table to open the Variant tab. Then click on Genotype Frequency from the menu on the left hand side of the page.

The most frequent allele at this position is C with a frequency of 0.850.

(d) Click on Sample genotypes in the left menu.

The table shows that sample 909454 has the C|T genotype and 909455 has the T|T genotype.