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Forensic Sciences (DNA Fingerprinting) STR Typing - Case Report
The document presents a forensic case report involving a badly burned female corpse, with key elements including the use of forensic entomology and DNA analysis for victim identification. The methodology described includes the extraction of DNA from maggots and buccal samples from an alleged father, alongside various DNA fingerprinting techniques such as PCR and capillary electrophoresis. The findings suggest a high probability of paternity testing and highlight the significance of entomological evidence in forensic investigations.
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Forensic Sciences (DNA Fingerprinting) STR Typing - Case Report
- 2. O Rabb! Openmy heart, ease my task, and loosen the knot from my tongue, so that they might fully understand my speech (Al TAHA).
- 4.
- 5. Case Report : •Badlyburned body --- Corpse obtained. •Face & neck colonized ----- Fly Larvae. •Soft Tissues ------Not available. •Feet & hands ----Missing. •Gender discrimination ----Not possible. •Small fraction of ----Burnt Liver. •Burned Bones. •Only Evidence ---Graduation Ring. •Woman --- Reported Missing (10 days earlier). •Genetic Profile --- Fragment of liver--- Unsuccessful. •Alleged Father.
- 6. Forensic Genetic Analysis Possiblesources of DNA • Blood • Semen • Saliva • Urine • Feces • Perspiration • Bone • Tissue
- 7. Forensic Entomology: Forensic entomologyis the application and study of insect and other arthropod biology to criminal matters.
- 8. Postmortem Interval: Time b/wthe death & discovery of the corpse.
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- 10. Working with evidences: • Fly Larvae – Maggots – Burnt Body. • Alleged Father. Procedure • DNA Finger Printing • Amelogenin PCR • Data Interpretation
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- 13. DNA from LarvaeGut was extracted by Following Procedure :
- 14. DNA from allegedFather • Buccal Sample collected. • DNA extracted by Chelex protocol. Five µl Buccal Sample + 200 µl of 5% Chelex 100. Highly-purified control DNA extracted with phenol- chloroform.
- 15. Southern blot 1. Restrictionenzyme digestion 2. Agarose gel electrophoresis 3. Transfer DNA onto membrane 4. Hybridized with probe RFLP Restriction Enzyme site 1. PCR amplify using flanking fluorescent primers 2. Capillary electrophoresis RFLP-based DNA fingerprinting PCR-STR based DNA fingerprinting PCR product of different sizes Fluorescent peaks of different mobility Radioactive bands of different mobility
- 16. PCR Multiplex kit ---AMPFistr Identifiler Kit Contents: •Allelic Ladders for Genotyping •PCR Component Mix •Primer Mix •Positive Control DNA Sample
- 18. CAPILLARY ELECTROPHORESIS ABI PRISM®310 Genetic Analyzer Automated gel pouring Automated sample injection Capillary electrophoresis with multi-color detection capabilities
- 19. ABI Prism 310Genetic Analyzer capillary Syringe with polymer solution Autosampler trayOutlet buffer Injection electrode Inlet buffer
- 20. D8S1179 D21S11 D7S820CSF1PO D3S1358 TH01 D13S317 D16S539 D2S1338 D19S433 D18S51 TPOX VWA AMEL D5S818 FGA GS500 LIZ size standard 6FAM (blue) VIC (green) NED (yellow) PET (red) LIZ (orange) AmpFlSTR® Identifiler™
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- 23. Repetitive sequence withinintron • Short repeated DNA sequence within intron • Present in different chromosomal locations • Unknown functions Minisatellites or microsatellite Exon Exon ExonIntron Intron
- 24. Mini. versus microsatellite •No. of repeats varies among individuals – VNTR (Variable Number of Tandem Repeats) • Minisatellites • Microsatellites • Minisatellites – Repeat unit length – 6-100 bases – Two to several hundred repeats at each minisatellite • Microsatellites – Repeat unit length: 1-7 bases – 5 to 100 repeats at each microsatellite – Thousands of different microsatellites, randomly scattered throughout genome http://www.wellcome.ac.uk/en/genome/genesandbody/hg07f006.html
- 25. Short Tandem Repeats(STR) • Each individual’s repeats is polymorphic in length – For example: • 25 % of population have 56 repeats in an STR • 50 % have 58 repeats • 25 % have 60 repeats – Repeats can be 2 – 7 bp • Repeat length at different STRs used as fingerprint. – Universal: can be used to amplify DNA from any individual
- 26. The CODIS System •National DNA Databank: CODIS • CODIS = Combined DNA Index System • In October 1998 the FBI's National DNA Index System (NDIS) became operational. • 13 CODIS Core STR Loci.
- 27. CSF1PO D5S818 D21S11 TH01 TPOX D13S317 D7S820 D16S539 D18S51 D8S1179 D3S1358 FGA VWA 13 CODISCore STR Loci AMEL AMEL Sex-typing Position of Forensic STR Markers on Human Chromosomes Penta E Penta D D2S1338 D19S433
- 28. STR Marker #AllelesRandom match probability (FBI Caucasian) CSF1PO 11 0.112 FGA 19 0.036 TH01 7 0.081 TPOX 7 0.195 VWA 10 0.062 D3S1358 10 0.075 D5S818 10 0.158 D7S820 11 0.065 D8S1179 10 0.067 D13S317 8 0.085 D16S539 8 0.089 D18S51 15 0.028 D21S11 20 0.039 Product 0.000000000000001683 One in 594,059,679,247,540 1 in 594 trillion Probability of a Random Match Using 13 CODIS STR Markers
- 30. Why STRs arePreferred Genetic Markers ? • Rapid processing is attainable • Abundant throughout the genome • Highly variable within various populations • Small size range allows multiplex development • Discrete alleles allow digital record of data • Allelic ladders simplify interpretation • PCR allows use of small amounts of DNA material • Small product size compatible with degraded DNA
- 32. Findings : • AmelogeninPCR ---- Female Corpse • Probability of paternity testing ----- 99.685 % • Results were later verified ----- DNA analysis of bone, possible after many attempts.
- 33. Discussion: • Accuracy • Reliability •Generation of good quality DNA • Storage of Maggots • Each maggot was processed separately, same results obtained • First reported case on forensic entomology in Mexico for victim identification.
- 34. References • Molecular SearchingTechniques http://tinyurl.com/bfbeal • DNA Diagnostics Center http://www.forensicdnacenter.com/dna-str.html • Brief History of Forensic DNA Typing http://tinyurl.com/dmbgvl • Wikipedia (easy reading for general principles) http://en.wikipedia.org/wiki/DNA_fingerprinting • CODIS (FBI) http://www.fbi.gov/hq/lab/codis/index1.htm • DNA fingerprinting protocols (very detailed) by Applied Biosystems and population genetics http://docs.appliedbiosystems.com/pebiodocs/04323291.pdf • Southern blot Flash http://www.pbs.org/wgbh/nova/sheppard/labwave.html • PCR principle http://www.wellcome.ac.uk/en/genome/technologies/hg17b009.html • http://www.cstl.nist.gov/biotech/strbase/
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Editor's Notes
- #7 No sources available in this case.
- #8 FE is the analysis of insect evidence for forensic & legal purpose.
- #12 Maggots collected in Microcentrifuge tube and preserved in 70 % ethanol at 4 C. Maggots taken from 3 diff. parts of the body. Processed separately. Larvae was washed , dissected and crop was removed from gut from which DNA was obtained.
- #16 RFLP-basedOriginal method developed by Alec JeffreysMinisatellite length polymorphism detected by RFLPBanding patternInvolves Southern blotMultiple steps; time consumingDifficult to automateDegradation of DNA may interfere fingerprint patternRequires relatively more DNAPCR-basedMicrosatellite length polymorphism detected by PCR product sizeFewer steps; quickAutomated PCR and capillary electrophoresisLess interfered by DNA degradationRequires less DNAStandard methods in crime labs including Hong Kong Government Laboratory