repetitive DNA Flashcards
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the human genome
what two things are repetitive DNA composed of
tandem repeats
interspersed repeats
what are tandem repeats composed of
Satellites
Minisatellites
Microsatellites – STR’s (short tandem repeats), SSRs (simple sequence repeats)
what are interspersed repeats composed of
LINES
SINES
LTRs
Transposons
what is nearly half of the human genome derived from
transposable elements
what are transposable elements
abundant in gene sequences and are also present in a significant portion of mature mRNAs, mostly in their untranslated regions
transposable elements are fundamental components of what
most eukaryotic genomes, with important contributions to their size, structure and variation. They occur as families with a variable copy number and they are (or were) able to transpose from one locus to another.
what are Class I elements also called
retroelements
what can Class I elements be divided into
LTR retrotransposons
what are LTR retrotransposons
flanked by long terminal repeats (LTRs) sharing an over-all organization similar to retroviruses and non-LTR retrotransposons, which terminate at one end with a poly(A) tract and have structural features of long and short interspersed nuclear elements (LINEs and SINEs, respectively).
what 2 things have LTR retrotransposons been further classified as
gypsy/Ty3-like
copia/Ty1-like
elements based on the order of their encoded proteins, that include a reverse transcriptase (RT) and integrase (IN).
what are Class II elements also called
(DNA) transposons
how are DNA transposons characterised
by terminal inverted repeats (TIRs)
how are DNA transposons grouped
into superfamilies on the basis of the target site duplication, the presence or not of the DDE triad (acidic amino acid motif), the sequence similarities at the DNA and protein levels (e.g. Tc1/mariner, hAT, Mutator-like, CACTA).
Most DNA transposons are organised in families of what two things
autonomous and non autonomous elements, characterized by their ability to respond to the same transposase.
satellite DNA
α (alphoid) 171bp, centromeres
β (Sau3A) 68bp, certain centromeres
Sat 1 25-48bp most centromeres
Sat 2 & 3 5bp all chromosomes
mini satellite DNA
Telomeric >6bp all telomeres
Hypervariable 9-64 many near telomeres
micro-satellite DNA
SSRs <10bp (or <6bp), dispersed
who discovered Hypervariable minisatellite DNA
Alec Jeffreys
what are Hypervariable minisatellite DNA
repetitive sequences within the DNA.
what varies between individuals of the mini satellite regions
Composition and length
what may be within these mini satellite regions
interspersed polymorphic restriction sites
what happens when you treat the DNA with endonucleases
cleaves the DNA into fragments to create a pattern that is unique to each individual.
how can the patterns of restriction fragments be viewed
Using the Southern blotting technique
This results in a barcode-like pattern which is unique to the individual. This became known as DNA fingerprinting.
advances to DNA fingerprinting
provides a simple pattern with high discriminatory power.
However, requires a large amount of sample material for the analyses, which is a disadvantage in criminal applications. In addition, inter-laboratory standardisation has been difficult to achieve and its relatively slow and expensive and difficult.
PCR of STR loci has overcome these difficulties and has greatly enhanced the role of DNA profiling in forensic science.
where are STRs found
dispersed throughout the whole genome and can be found in large numbers, even in the gene introns.
what are the most common STRs
Dinucleotide polymorphisms but they are strongly affected by strand slippage during amplification, producing artefactual stutter bands
what STR repeats are less prone to slipping
Tetra- and pentanucleotide repeats so more suitable for forensic analysis
STR methods
can be easily standardised and are suitable for multiplexing. Amplicon (PCR product) length measurement is automated; its cheap, quick, accurate and easy.
what are STRs, SSRs or micro satellites defined as
tandemly repeated tracts of DNA composed of 1–6 base pair long motifs.
The number of repeats in a tract is usually less than ten and if it is higher then repeat length polymorphism is often observed. In rare cases, the repeat number may reach hundreds and even thousands.
all SSRs together occupy how much of the human genome
3%
• At each STR locus there are two alleles and an individual will be either
- Homozygous – e.g. AGAT repeated 8 times, AGAT repeated 8 times, reported as 8,8 genotype
- Heterozygous – e.g. AGAT repeated 8 times, AGAT repeated 5 times, reported as 5,8 genotype
simple sequence repeats
- The genomic distribution of the SSRs of different motif lengths and sequences is strongly biased. With the exception of monomeric motifs, the classes of repeated dimers, tetramers and hexamers, which show nearly equal density in the genome, are 3–4 times more abundant than those of trimers and pentamers.
- When the occurrence of SSRs in different functional genome regions is considered, it turns out that most of them show much higher density in non-coding regions.
- Exceptions to the rule are trimers and hexamers which are nearly two times more prevalent in exons compared to introns and intergenic regions
microsatellites
- In the human (and mouse) genome, microsatellite density is nearly twofold higher near the ends of chromosome arms.
- Microsatellites are also frequently found in the proximity of interspersed repetitive elements such as short interspersed repeats (SINEs) and long interspersed elements (LINEs).
SGM+ is now being replaced with what
DNA-17
what is Amelogenin
not an STR
its a length polymorphism and used in sex determination
appearance of PCR products
• PCR products are small in size and products from different loci overlap – hence the need for a fluorescent label.
the more alleles at each locus…
the better discrimination providing allele frequencies are approximately equal.
point to note
not all STRs are simple motifs that perfectly repeat a tetra nucleotide motif
see pp for
image of replication slippage
see pp for
basic gene structure and expression
