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Applications of HGP Genetic testing Forensics. testing for a pathogenic mutation in a certain gene in an individual that indicate a persons risk of developing.

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Presentation on theme: "Applications of HGP Genetic testing Forensics. testing for a pathogenic mutation in a certain gene in an individual that indicate a persons risk of developing."— Presentation transcript:

1 Applications of HGP Genetic testing Forensics

2 testing for a pathogenic mutation in a certain gene in an individual that indicate a persons risk of developing or transmitting a disease Used for mutation screening of disease genes e.g. HD, CFTR, DMD Genetic testing

3 Directly Gene tracking Population screening Genetic testing can be done either

4 DIRECT GENETIC TESTING Based on either a)MUTATION DETECTION: screening for KNOWN polymorphisms in DNA b)MUTATION SCANNING: screening for UNKNOWN polymorphisms in DNA

5 SNPs by RFLP-PCR Must have sequence on either side of polymorphism –Amplify fragment –Expose to restriction enzyme –Gel electrophoresis e.g., sickle-cell genotyping with a PCR based protocol Fig Hartwell MUTATION DETECTION

6 Very short specific probes (<21 bp) which hybridize to one allele or other Such probes are allele-specific oligonucleotides (ASOs) Fig SNPs by ASOs MUTATION DETECTION

7 Variation in length of DNA sequence (repetitive DNA) pathogenic (Huntingtons disease) non pathogenic (forensics) MUTATION DETECTION classSize of repeat Repeat block Major chromosomal location minisatellite9-64 bp0.1 – 20kbTelomeres microsatellites1-13 bp< 150 bpDispersed

8 Huntingtons disease -a microsatellite triplet repeat in a coding region Figure 18.12: HMG3


10 METHODS MUTATION SCANNING Direct sequencing Southern blots dHPLC Microarrays etc sequencing

11 MUTATION SCANNING Using dHPLC Exon 6 of DMD gene normal affected Fig18.4: HMG3 by Strachan & Read

12 MUTATION SCANNING Using multiplex ARMS test Screening for 29 mutations of the CFTR gene Fig18.10: HMG3 by Strachan & Read

13 GENE TRACKING Analysis of linked markers in families for the inheritance of a high risk chromosome from heterozygous parents. The process has 3 steps 1) find a closely linked marker for which the parents are heterozygous 2) work out which chromosome carries the disease allele 3) work out which chromosome the individual has inherited Used when map location of disease locus is known but not the actual disease gene

14 POPULATION SCREENING Screening programs for well characterised traits must be both SENSITIVE ACCURATE e.g. PKU tests /Guthrie (PAH activity) ARMS test (CFTR mutations)

15 Forensics Identify crime suspects / exonerate innocent Identify victims Establish family relationships Identify endangered species Detect pollutants Match organ donor with recipient Determine seed / livestock pedigree Authenticate consummables

16 Extract DNA Analyse specific regions using probes look for matches between 2 samples at many loci (multilocus) Scan ~ 10 DNA regions that show locus variability > 5 matches Create DNA profile (DNA fingerprint) How does forensic ID work?

17 DNA fingerprinting Originally described using minisatellite probes consisting of tandem repeats of the myoglobin locus ( Nature, 1985, 316: Jeffereys et al) Number of multiple loci probes (MLP) identified Core sequence GGAGGTGGGCAGGA 2 of these used (33.15 and 33.6) Together, upto 36 independently inherited bands detected


19 DNA fingerprinting superceded by single locus probes (SLP) – just 2 bands per probe Now superceded by SL-PCR Use of allelic ladder markers Advantages Increased sensitivity Small sample quantities sufficient Uses microsatellites instead of minisatellites

20 Simple sequence repeats (SSRs) Microsatellites 1-13 bp repeats e.g. (A) n (AC) n Minisatellites bp repeats 3% of genome (dinucleotides - 0.5%) Repetitive sequences… HUMFES/FPS (ATTT) 8-14

21 DNA fingerprinting 1995 – National Criminal Intelligence Database (Forensic science service) 700,000 samples stored Strength of evidence based on likelihood ratio (LR) LR = C / C PROSECUTORS FALLACY The probability of the DNA evidence, if it came from the suspect, is 1 in 50 million


23 DNA fingerprints can identify individuals and determine parentage E.g., DNA fingerprints confirmed Dolly the sheep was cloned from an adult udder cell Donor udder (U), cell culture from udder (C), Dollys blood cell DNA (D), and control sheep 1-12 Fig Hartwell

24 Is DNA effective in identification? Only if used intelligently!! Only regions showing the most variability must be used Must cover large regions Look for matches beyond a reasonable doubt

25 Mitochondrial DNA Multicopy 16.5 kbp Maternally inherited Sequenced in 1981 (Nature,1981, 290:457-65) Mutation rate ~1/33 generations Heteroplasmy (original and mutated forms co- exist) More stable for forensic analysis

26 Mitochondrial DNA Highest variation in control region (800bp)

27 Y chromosome Haploid Paternal inheritance Binary polymorphisms

28 References Hum Mol Gen 3 by Strachan and Read Chapter 18 Hartwell et al – Chapter 11; pages DNA profiling in forensics by Peter Gill et al

29 Bioarchaeology, Anthropology, Evolution, and Human Migration study evolution through germline mutations in lineages study migration of different population groups based on female genetic inheritance study mutations on the Y chromosome to trace lineage and migration of males compare breakpoints in the evolution of mutations with ages of populations and historical events Applications of HGP

30 Microbial Genomics new energy sources (biofuels) environmental monitoring to detect pollutants protection from biological and chemical warfare safe, efficient toxic waste cleanup understanding disease vulnerabilities and revealing drug targets Applications of HGP

31 Risk Assessment assess health damage and risks caused by radiation exposure, including low-dose exposures assess health damage and risks caused by exposure to mutagens & carcinogens reduce the likelihood of heritable mutations Applications of HGP

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