1. Lecture 2 – Repeat elements
BB30055: Genes and genomes
Genomes - Dr. MV Hejmadi
Lecture 2 – Repeat elements

2. Lecture 2 Repeat elements
What are repeat elements?
How did they originate?
Why are they important?

4. Repetitive DNA Main classes based on origin Tandem repeats
Interspersed repeats
Segmental duplications

5. 1) Tandem repeats/ clustered repeats
Blocks of tandem repeats at
subtelomeres
pericentromeres
Short arms of acrocentric chromosomes
Ribosomal gene clusters

6. Tandem / clustered repeats
Broadly divided into 3 types based on size and location
class
Size of repeat
Repeat block
Major chromosomal location
Satellite
5-171 bp
> 100kb
centromeric
heterochromatin
minisatellite
9-64 bp
0.1–20kb
Telomeres
microsatellites
1-13 bp
< 150 bp
Dispersed
Which of the classes of tandem repeats are likely to be found in a mRNA and which are unlikely?
HMG3 – Chap 9 pp

7. Satellites Large arrays of repeats Some examples Satellite 1,2 & 3
Sat a (Alphoid DNA)
- found in all chromosomes
Sat b satellite
Coutesy: Evani Viegas-Pequignot
Alphoid contains centromere binding protein CENPB
ICF syndrome
(Immunodeficiency- Centromeric instability-Facial anomalies syndrome)
Caused by hypomethylation of Sata in chromosomes 1,9,16

8. Minisatellites Moderate sized arrays of repeats Some examples
Hypervariable minisatellite DNA
- core of GGGCAGGAXG
- found in telomeric regions
- used in original DNA fingerprinting technique by Alec Jeffreys
Similar to ecoli chi sequence – hotspot for recombination

9. Microsatellites VNTRs - Variable Number of Tandem Repeats,
SSR - Simple Sequence Repeats /STR – short tandem repeats
1-13 bp repeats e.g. (A)n ; (AC)n
2% of genome (dinucleotides - 0.5%)
Used as genetic markers (especially for disease mapping)
A father might have a genotype of 12 repeats and 19 repeats, a mother might have 18 repeats and 15 repeats while their first born might have repeats of 12 and 15.

10. Microsatellite genotyping
design PCR primers unique to one locus in the genome
a single pair of PCR primers will produce different sized products for each of the different length microsatellites .

11. Genotype test in a large family using (CA)/(TG) marker D17S800
1 (3,6)
2 (1,5)
3 (3,5)
4 (2,5)
5 (3,6) etc
Alleles
Samples
What is the genotype of sample 10, 13, 16 ?
Why do you get the shadow bands?
Suggested to be due to replication slippage

12. strand slippage during replication
How are tandem repeats generated in the genome?
strand slippage during replication
Fig 11.5 HMG3 by Strachan and Read pp 330

13. strand slippage during replication

14. 2) Interspersed repeats
A.k.a. Transposon-derived repeats
~ 45% of genome
Arise mainly as a result of
transposition either through DNA or RNA
retrotransposons (retroposons)
‘copy and paste’
DNA transposons (‘cut & paste’)
See lecture 3 for transposition

15. Classes of transposable elements
Classes of mobile elements. DNA transposons, e.g., Tc-1/mariner, have inverted terminal inverted repeats (ITRs) and a single open reading frame (ORF) that encodes a transposase. They are flanked by short direct repeats (DRs). Retrotransposons are divided into autonomous and nonautonomous classes depending on whether they have ORFs that encode proteins required for retrotransposition. Common autonomous retrotransposons are (i) LTRs or (ii) non-LTRs (see text for a discussion of other retrotransposons that do not fall into either class). Examples of LTR retrotransposons are human endogenous retroviruses (HERV) (shown) and various Ty elements of S. cerevisiae (not shown). These elements have terminal LTRs and slightly overlapping ORFs for their group-specific antigen (gag), protease (prt), polymerase (pol), and envelope (env) genes. They produce target site duplications (TSDs) upon insertion. Also shown are the reverse transcriptase (RT) and endonuclease (EN) domains. Other LTR retrotransposons that are responsible for most mobile-element insertions in mice are the intracisternal A-particles (IAPs), early transposons (Etns), and mammalian LTR-retrotransposons (MaLRs). These elements are not present in humans, and essentially all are defective, so the source of their RT in trans remains unknown. L1 is an example of a non-LTR retrotransposon. L1s consist of a 5'-untranslated region (5'UTR) containing an internal promoter, two ORFs, a 3'UTR, and a poly(A) signal followed by a poly(A) tail (An). L1s are usually flanked by 7- to 20-bp target site duplications (TSDs). The RT, EN, and a conserved cysteine-rich domain (C) are shown. An Alu element is an example of a nonautonomous retrotransposon. Alus contain two similar monomers, the left (L) and the right (R), and end in a poly(A) tail. Approximate full-length element sizes are given in parentheses
Science 12 March 2004: Vol no. 5664, pp

16. Interspersed repeats (transposon-derived)
major types
class
family
size
Copy number
% genome*
LINE
L1 (Kpn family) L2
~6.4kb
0.5x106
0.3 x 106
16.9
3.2
SINE
Alu
~0.3kb
1.1x106
10.6
LTR
e.g.HERV
~1.3kb
0.3x106
8.3
DNA
transposon
mariner
~0.25kb
1-2x104
2.8
* Updated from HGP publications
HMG3 by Strachan & Read pp

17. Repeat elements in the human genome

18. 3) Segmental duplications
Closely related sequence blocks (1-200kb) at different genomic loci
Segmental duplications can occur on homologous chromosomes (intrachromosomal) or non homologous chromosomes (interchromosomal)
Not always tandemly arranged
Relatively recent

19. Segmental duplications
Segmental duplications in chromosome 22

20. Repeat elements How did they originate?
Tandem repeats – replication slippage etc
Interspersed repeats – transposition events
Segmental duplications – strand exchange, recombination events

21. Repeat elements Why are they important? Evolutionary ‘signposts’
Passive markers for mutation assays
Actively reorganise gene organisation by creating, shuffling or modifying existing genes
Provides information on chromosome structure and dynamics
Provides tools for medical, forensic, genetic analysis

22. Pathogenic potential of Short Tandem Repeats (STR)
Reduction or expansion of STR can be pathogenic
1) Unstable expansion of short tandem repeats
Characterised by anticipation
Large expansions outside coding sequences
Modest expansions within coding sequences
FRAXA, FRAX E
Huntington disease (HD)
Myotonic dystrophy (DM1)
SCA 1,2,3,6,7, 17
Friedrich ataxia (FA)
Kennedy disease
Spinocerebellar ataxia 8,11
Anticipation – lower age of onset and/or severity is worse over subsequent generations
FA – Freidrich’s ataxia HD
10-30 CAG repeats – non pathogenic
repeats – unstable repeats cause huntingtin protein to aggregate and kill cells
Pg 477 HMG3
HMG 3 Chapter 11 pp

23. 2) Unstable deletions of STRs
STRs tend to be deletion hotspots
Pg 358-HMG3

24. Interspersed repeats are susceptible to deletions/duplications
E.g. Kearns-Sayre syndrome- encephalomyopathy
External opthalmoplegia
Ptosis
Ataxia
Cataract
Common 4977bp deletion in mt DNA
External opthalmoplegia
Ptosis
Ataxia
Cataract
Mt is recombination deficient therefore such deletions could arise due to replication slippage just like the STR

25. Pathogenic potential of segmental duplications
Nature Reviews Genetics 2, (2001)

26. STRs are… 
A) microsatellite sequences.
B) generally highly polymorphic.
C) Useful markers along the chromosome.
D) a and b
b and c
a and c

27. Which of the following is not one of the four classes of transposon-derived repeat sequences? 
A. LINES
B. SINES
C. DNA transposons
D. Long terminal repeat transposons
E. pseudogenes

28. When repeats of a region lie adjacent to each other they are called ___________ duplications
________________ gene transfer is the direct transfer of genes from one species in the germ line of another species.

29. True or False
All Repetitive DNA elements in the human genome arose through a common mechanism

30. Match the columns Recombination events Tandem repeats
Replication slippage
Transposition events
Tandem repeats
Interspersed repeats
Segmental duplications

31. Reading Chapter 9: HMG 3 by Strachan & Read
Repeat elements and disease
Nature Reviews Genetics 2, (2001)

32. CpG islands show no methylation
TpG
Methyl CpG
Deamination
methylated at C
CpG islands show no methylation
Significance of CpG islands
Non-methylated CpG islands associated with the 5’ ends of genes
Usually overlap the promoter region
Aberrant methylation of CpG islands linked to pathologies like cancer or epigenetic diseases like Rhett’s syndrome

33. Inheritance of CpG methylation