Thursday, February 19, 2015

GRC Website: Individual Genome Issues

Are you looking for the latest status updates from the GRC on the human, mouse or zebrafish reference genome assemblies? In companion to our previous post, we now explain how to use the Individual Genome Issue reports on the GRC website. As described in our last blog, you can filter and search for issues of interest using the organism-specific "Issues Under Review" pages. To provide an example for this blog post, we applied the follow filtering options on the human "Issues Under Review" page: issues location = GRCh38.p2, chromosome = chr3 , type = variation and scaffold type = ALT (alternative loci). We then selected HG-1291 from column 1 of the results table to go to the individual issue page, shown below in Figure 1. On this and other individual issue pages, you'll find the following information:
  • Summary fields describing the issue and its latest status updates or resolution (blue box)
  • Ideogram showing the issue's genomic location (green box)
  • Patch and/or alternate loci status and history (orange box)
  • Graphical view of genomic region to which issue is mapped (red box). 
    • Note: graphical views are provided for all mapped locations in the previous and current assembly versions. For example, HG-1291 has been mapped to chr. 3 and an alternate locus scaffold in GRCh38.p2, and to chr. 3 and a novel patch in GRCh37.p13. Use the radio buttons to toggle the display between the different sequence locations.
Figure 1. GRC Issue page for HG-1291, with page features highlighted.

Below, figure 2 shows the graphical view of the GRCh38.p2 alternate locus scaffold to which HG-1291 has been mapped (NW_003871060.2). Default tracks in the graphical views provide you with additional information about the assembly composition and quality. They include:

  • Assembly components
  • Alignments of alternate loci/patch scaffolds to the primary assembly
  • Annotated component assembly problems
  • All GRC issues mapped to the region
  • NCBI Gene annotation
  • Ensembl Gene annotation

In this image of HG-1291, review of the Genes and Alignment tracks reveals two exons in a region of the alternate loci that has no alignment to the chromosome (arrow and circle). This annotation supports the description in the Issue Summary fields. You can further configure the tracks or upload your own data files to the graphical view by clicking on the "Configure" button at the top right of the viewer (red box).
Figure 2. Graphical view of NW_003871060.2, the GRCh38.p2 alternate loci scaffold to which issue HG-1291 is mapped. The exons captured by the additional sequence in the scaffold are highlighted.

If you have questions about any of the issues you see, please contact the GRC and reference the issue number. If you know of a genome issue that isn't found on these pages, please report the issue to the GRC.

Tuesday, February 3, 2015

GRC Website Update: Genome Issues Under Review

GRC "Genome Issues under Review" webpage update!

Do you know how to find genome issues on the GRC website? To get started, select an organism from the top of the GRC homepage, and in the corresponding organism overview page select the link for "Issues Under Review". These pages provide you with the latest information about potential problems and other issues related to the human, mouse and zebrafish reference genome assemblies that the GRC are working on. Recent updates to these pages make them more interactive, informative and easier to navigate so you can pinpoint issues relevant to your research interests. Some of the page features are highlighted in Figure 1, which shows "Human Genome Issues".
  • Show issue locations on (blue box): Use this to define the assembly version on which you want to see mapped issues. We support issue mapping to the current assembly and the last release of the prior assembly version.
  • Ideogram (green box): The histogram above presents the number of issues related to each chromosome, and the annotations show issue locations. Looking for issues related to a single chromosome? Click on a chromosome or histogram of interest to see a more detailed ideogram with annotated issues (more on this below).
  • Search (purple box)Use this to finding issues related to a specific gene/clone/accession number/chromosomal location.
  • Data table: Provides a summary of issues. Within this table, click on issue ID (brown box) to go to web pages for specific issues or View in browsers (brown box) to see the relevant genome regions in browsers at Ensembl, NCBI, and UCSC.
Figure 1. Human Genome Issues overview
Additional page features shown below in Figure 2 will help you identify the issues that interest you most:
  • Filter: Located to the left of the data table, this section contains various display filters, including issue type and issue status, to help you find GRC issues meeting specified criteria.
  • Issue Annotations: In the single chromosome ideogram displays, issues are annotated below the figure.
    • Tool-tips: Click on any annotation for a summary and a link to the issue page
    • Bar charts: Click on either of the interactive bar charts below the ideogram to re-categorize the issue annotation display by Type or Status.
Figure 2. Chromosome 1 genome issues
If you have questions about any of the issues you see, please contact the GRC and reference the issue number. If you know of a genome issue that isn't found on these pages, please report the issue to the GRC.

Wednesday, January 28, 2015

GRCh38: Patching the ABO gene

GRCh38 has started receiving patch updates, and this blog post describes a FIX patch to the ABO gene, located on chr. 9. You might have been aware that the GRC released a FIX patch to ABO for GRCh37. So why is there an ABO FIX patch for GRCh38 as well?

In GRCh37, the ABO gene was annotated on sequence derived from two RP11 library clones, AL732364.9 (RP11-244N20) and AL158826.23 (RP11-430N14). However, the RP11 library is derived from a diploid genome and analysis demonstrated that the two sequenced clones represented two different Type O ABO alleles. As a result, the GRCh37 chr.9  representation of ABO was an invalid haplotype for the gene (Fig. 1, top panel).
Fig. 1 Top: ABO region in GRCh37. The gene is derived from 2 components, resulting in an invalid  haplotype not seen in any individuals. Bottom: ABO fix patch. The gene is derived from a single component and represents a known Type O haplotype.

To address this issue, we identified a clone from the CalTech human BAC library D that captures the complete ABO gene (CTD-2612A24). The sequence for this clone was finished (AL772161.10) and inserted into the chr. 9 tiling path, replacing RP11 component AL158826.23. By setting the switch points between AL732364.9 (RP11-244N20) and AL772161.10 (CTD-2612A24) so that the full insert sequence of the new component contributed to the scaffold, we were able to provide a complete and valid ABO Type A1.02 representation for the gene. Thus update was provided as a FIX patch scaffold (GL339450.1) for GRCh37. (Fig.1, bottom panel).

Unfortunately, this update is not reflected in GRCh38. Subsequent to the final GRCh37 patch release (GRCh37.p13) and the release of GRCh38, the sequence to RP11-244N20 was updated (AL732364.10) and inserted into the chr. 9 tiling path. The switch points between the updated sequence AL732364.10 and AL772161.10 were set incorrectly (Fig. 2). This resulted in an invalid haplotypic representation for ABO. Whereas the GRCh37 representation was a Type O/O mix, in GRCh38 it is a Type A/O mix.
Fig.2 Top: ABO fix patch. Gene is derived from a single component. Bottom: ABO region in GRCh38. The gene is derived from 2 components, creating an invalid haplotype. This is fixed by the GRCh38 FIX patch.

The GRCh38 FIX patch scaffold KN196479.1 corrects this switch point and provides the same single haplotype representation for ABO that was present in the GRCh37 FIX patch scaffold. This re-patching of the ABO gene again restores the functionality of the gene with the valid Type A1.02 haplotype.

Thursday, November 27, 2014

Optical Mapping data in the GRC track hub

The GRC track hub now includes Optical Mapping analysis information.

What is Optical Mapping?
Optical Mapping (OM) is a method to produce ordered restriction maps from single DNA molecules (rMaps).  These rMaps are assembled into consensus maps which can be aligned against the reference assembly, taking into account the positioning of restriction sites and length of fragments. OM aids the scaffolding of genomic sequence and the identification of errors in genome assemblies, but it is also very helpful in confirming assembled contigs and sizing gaps.

What OM data is available?
OM data is currently available for human (Teague et al., 2010) and mouse (Church et al., 2009) and we would like to thank Steve Goldstein and David Schwartz for providing the alignments to the respective reference assemblies.

What is displayed in the GRC track hub?
The OM data is divided into several tracks in the GRC track hub. These tracks are of three types:
  • OM alignment tracks show the alignments of consensus maps to the reference genome, based on the comparison of restriction patterns. Each track of this type corresponds to an analysis of OM data from a single cell line.
  • OM deletion tracks present the locations of additional restriction fragments that have no corresponding fragment in the reference assembly. Their position is defined by the remaining alignment of the respective consensus map. Again, each track of this type corresponds to an analysis of OM data from a single cell line.
  • Each assembly also has a single OM reference track, which presents the set of OM fragments that would be expected based on the reference sequence, produced via an in silico restriction digest.
How is this information visualised?
The way that OM analysis data is displayed is slightly different for each of the types of track mentioned above (alignments, deletions, and predicted fragments based on the reference).
  • The OM alignments tracks present each contig as a horizontal line, with restriction cut-sites dividing fragments being displayed as vertical lines along that contig. Where there is a space between the placement of successive restriction fragments according to this analysis, this is represented as a thicker vertical bar spanning the gap between the fragments.
  • The OM deletions tracks use a single vertical bar to show the location of each fragment or group of fragments with no corresponding fragment in the reference assembly. The size of the fragment is not represented by the glyph in the browser, but is shown as one of its data fields.
  • The OM reference track displays the cut-sites between expected restriction digest fragments as vertical lines.
Display examples
In the Ensembl genome browser (from version 78 onwards), the OM reference track is at the top, with the OM deletions track "OM gap 15510" below it, followed by three OM alignments tracks based on different cell lines. (Note that OM deletions tracks exist for all those cell lines which have alignments tracks, but only one OM deletion track has data at this location.)

Here is how it appears in the UCSC genome browser. The tracks are in the same order as for the Ensembl example above: the OM reference track is at the top, with the OM deletions track "OM gap 15510" below it, followed by three OM alignments tracks based on different cell lines.

    Tuesday, October 21, 2014

    GRC track hub arrives!

    The GRC are now providing assembly-related tracks for reference genomes via a track hub, which will allow you to view that data in a range of genome browsers, including Ensembl and UCSC. The location of this hub is:

    What tracks are available?
    The GRC generates a range of annotations on the reference genomes it curates. These tracks describe the assembly of the genome, as well as quality issues with the genome, and provides information relevant to their resolution or planned improvement. Much of this annotation is already available via the gEVAL browser. However, the track hub allows you to view this annotation in the genome browser of your choice.

    The individual tracks currently available are:
    • Genome issues under review by the GRC
    • Genomic regions defined by the GRC
    • Alignments between the primary assembly and alternate loci or patches
    • Clone sequence anomalies
    • Human regions with clones from the CHORI-17 library (CHM1tert)
    These tracks are updated on a weekly basis. We will be adding to this range of information as time goes on.

    How does this look?
    Here's how this data looks in Ensembl:
     Here's how it looks in UCSC:

    What is a track hub?
    A track hub is a means of attaching multiple annotation tracks to a genome browser via a single URL. Full documentation on track hubs is available here.

    How do I attach the track hub in my favourite genome browser?
    You need to specify the following URL to the genome browser:
    • In Ensembl: Go to "Add/Manage your data", select "Add your data" (if necessary), then select the data format "TrackHub", and add the hub URL.
    • In UCSC: select the "Track Hubs" button just beneath the main browser area, then select the "My Hubs" tab, and add the hub URL.

    Tuesday, October 14, 2014

    GRCh38.p1 has arrived!

    The first patch release for the GRCh38 reference assembly is now available. The GRCh38.p1 release includes 16 scaffolds: 13 FIX patches and 3 NOVEL patches. The FIX patch scaffolds correct existing assembly sequences, while the NOVEL patch scaffolds provide new alternate sequence representations. You can download the GRCh38.p1 assembly, including the alignments of the patches to GRCh38, from the GenBank FTP site:

    Stay tuned for upcoming blog posts on individual patches!

    Wednesday, September 17, 2014

    GRCz10 - The GRC's first zebrafish genome reference assembly

    When the Zv9 assembly was released in July 2010, the zebrafish genome sequence was given into the care of the GRC for future improvement and maintenance. After 4 years of hard work (and zebrafish IS hard work), we have now produced a new reference assembly, GRCz10.

    The previous assembly was already of high value for the scientific community, and served well for both the investigation of isolated gene loci and to address overall bioinformatics questions (Howe et al. 2013), but still featured many gaps and suffered from sub-optimal long-range continuity. To address this, we have sequenced more than 1500 additional BAC and fosmid clones and added them to the assembly. We reviewed clone overlaps and clone placements with a variety of techniques. In collaboration with the Stemple lab, using the MGH panel, we generated a new meiotic map to fill remaining gaps in the high density meiotic map SATMAP. This new map, GAPMAP, helped with placing previously unlocalised contigs onto chromosomes, and allowed us to assess and improve the order of existing chromosome placements. The creation of an optical map further improved the clone assignments, with a notable impact on the structure of the repeat-rich chromosome 4. Thanks to a collaboration with Mark Hills from the Lansdorp lab, we gained additional insight into the orientation of assembly components, leading to more than 250 orientation changes and re-placements.  In total, more than 4000 genome issues were reviewed and resolved. The remaining gaps in the clone path were filled with sequence from the WGS31 whole genome assembly, as done before with Zv9.

    The most notable changes in the chromosome landscape since Zv9 can be found on chromosome 4, which has gained about 15 Mb in length, and 94 of the 112 previously unplaced clone-contigs found a home on a chromosome. Whilst 85% of all publicly available cDNAs could be assigned a place on Zv9 with at least 97% identity and 90% coverage, we now find 87% in GRCz10. If we classify cDNAs with less than 97% identity and less than 40% coverage as not found, then Zv9 was missing 7% of the cDNAs, whilst GRCz10 now is only missing 3%. Now that the assembly has been released, the Havana team at the Sanger Institute is busy manually (re-)annotating genes, and the Ensembl team is working on generating an automated gene build and integrating it with these manually produced models. The NCBI eukaryotic genome annotation pipeline (gpipe) will also annotate the GRCz10 RefSeq assembly.

    If you are working with the zebrafish genome assembly, we'd be very happy to get some feedback from you. You can either fill in the form at the GRC home page, or send us an email to