The document discusses the human reference genome assembly GRCh38. It provides information on assembly basics, the assembly model used, updates made in GRCh38 compared to the previous version, and how to access and utilize the sequence and annotation data. Key points include that GRCh38 represents 178 genomic regions with alternative loci sequences, contains over 400kb of novel sequence from targeted updates, and is collaboratively maintained to integrate new data.

1. Getting the Most from the Reference
Assembly
Valerie Schneider, Ph.D.
NCBI
6 October 2015
http://genomereference.org

2. http://genomereference.org
Twitter: @GenomeRef
grc-announce@ncbi.nlm.nih.gov

3. Outline
• Assembly basics
• The assembly model
• GRCh38 & updates
• Taking advantage of the data

4. Reference Assembly Basics

6. Sims et al. (2014) Nat Rev Genet. 15(2):121-32
30x
1x
Reference Assembly Basics
Lander and Waterman
(1988) Genomics SequencedNot sequenced
1X Coverage
5X Coverage
10X Coverage
37% 63%
0.6% 99.4%
0.005% 99.995%

7. Reference Assembly Basics
FINISHED?

8. BAC insert
BAC vector
Shotgun sequence
Assemble
Foldsequence
Gaps
deeper sequence
coverage rarely
resolves all gaps
GAPS
“finishers” go in to manually
fill the gaps, often by PCR
Clone based assemblies
Reference Assembly Basics
Minimal Tiling Path

9. Human assemblies available in the NCBI assembly database
http://www.ncbi.nlm.nih.gov/assembly
Reference Assembly Basics
Oct. 2014: 13 assemblies
Oct. 2015: 25 assemblies
YRI
CEU
CEU
CHB

10. Reference Assembly Basics
Sanger Sanger Illumina Illumina PacBio (older)
clone WGS WGS WGS WGS
Reads:
Method:
PacBio (newer)
WGS
N50:
Measure of continuity.
Half of the contigs in the
assembly are this length or
greater.Why all this matters:
Longer haplotype blocks
Fewer collapsed repeats & segmental duplications
Improved annotation
More robust mapping target

11. Outline
• Assembly basics
• The assembly model
• GRCh38 & updates
• Taking advantage of the data

12. Sequences from haplotype 1
Sequences from haplotype 2
Old Assembly model: compress into a consensus
Current Assembly model: represent both haplotypes
GRC Assembly Model
many

13. Assembly (e.g. GRCh38)
Primary
Assembly
Unit
Non-nuclear
assembly unit
(e.g. MT)
PAR
Genomic
Region
(MHC)
Genomic
Region
(UGT2B17)
Genomic
Region
(MAPT)
Church et al., PLoS Biol. 2011 Jul;9(7):e1001091
GRC Assembly Model
ALT
2
ALT
3
ALT
4
ALT
5
ALT
6
ALT
7
ALT
1

14. GRC Assembly Model
Alt loci alignments are an integral part of the assembly model
alignment to chr + scaffold sequence = Alt

15. GRCh38
• 178 regions with alt loci: 2% of chromosome
sequence (61.9 Mb)
• 261 Alt Loci: 3.6 Mb novel sequence relative to
chromosomes
• Average alt length = 400 kb, max = ~5 Mb
GRCh38

16. Outline
• Assembly basics
• The assembly model
• GRCh38 & updates
• Taking advantage of the data

17. GRCh38: Alt Loci
Alignment Legend
no alignmentmismatchdeletion

18. chromosome
alt/patch
reads On-target alignment
Off-target alignments
(n=122,922)
GRCh38: Alt Loci

19. GRCh38: Assembly Stats
http://genomereference.org
GRCh38 vs. GRCh37

20. GRCh38: Annotation Stats

21. GRCh38 Base Updates
Targeted PCR/WGS: n=91

22. GRCh38 Centromeres
Miga et al., Genome Res. 2014 Apr;24(4):697-707

23. GRCh38 Novel Sequence

24. GRCh38 Novel Sequence

25. Assembly (e.g. GRCh38.p1)
Primary
Assembly
Unit
Non-nuclear
assembly unit
(e.g. MT)
ALT
1
ALT
2
ALT
3
ALT
4
ALT
5
ALT
6
ALT
7
PAR
Genomic
Region
(MHC)
Genomic
Region
(UGT2B17)
Genomic
Region
(MAPT)
Patches
Genomic
Region
(ABO)
Genomic
Region
(FOXO6)
Genomic
Region
(FCGBP)
Assembly Updates
Patches
FIX NOVEL
SCAFFOLD STATUS AT NEXT
MAJOR ASSEMBLY RELEASE
ALT
LOCI
--
(integrated)
Treat as:
Allelic
Treat as:
Preferred

26. GRCh38.p4
• 55 Patches: >400 kb novel sequence
• 37 FIX
• 18 NOVEL
Assembly Updates
Learn more about
assembly updates at
the GRC poster:
1834W (6-7 pm)

27. Outline
• Assembly basics
• The assembly model
• GRCh38 & updates
• Taking advantage of the data

28. Accessing the Data
http://genomereference.org

29. Accessing the Data

30. Accessing the Data

31. Accessing the Data

32. Accessing the Data

33. Accessing the Data

34. GRC Assembly Management

35. Accessing the Data
http://www.ensembl.org/

36. Accessing the Data

37. Accessing the Data
ftp://ngs.sanger.ac.uk/production/grit/track_hub/hub.txt

38. Accessing the Data

39. http://www.ncbi.nlm.nih.gov/variation/view/NCBI Variation Viewer
Accessing the Data
Learn more about
viewing GRCh38 at
NCBI: 1748T (12-1 pm)

40. http://www.ncbi.nlm.nih.gov/genome/tools/remap

41. Outline
• Assembly basics
• The assembly model
• GRCh38 & updates
• Taking advantage of the data

42. GRCh38 Collaborators
• NCBI RefSeq and gpipe annotation team
• Havana annotators
• Karen Miga
• David Schwartz
• Steve Goldstein
• Mario Caceres
• Giulio Genovese
• Jeff Kidd
• Peter Lansdorp
• Mark Hills
• David Page
• Jim Knight
• Stephan Schuster
• 1000 Genomes
GRC SAB
• Rick Myers
• Granger Sutton
• Evan Eichler
• Jim Kent
• Roderic Guigo
• Carol Bult
• Derek Stemple
• Jan Korbel
• Liz Worthey
• Matthew Hurles
• Richard Gibbs
GRC Creditshttp://genomereference.org