HID Flashcards
(40 cards)
HID
- novel concept of DNA fingerprinting
- discovered by Sir Alec Jeffreys
- first published in 1985 and first forensically used in 1987
what are the common uses of HID
- forensic cases= identification of suspect
- paternity= identification or confirmation of father
- missing person= identification
- Disaster victim ID
- military dog tags
- criminal databases
The genome
every living thing has a genome
- contain biological info needed to construct you
- genome of forensic significance is made up of DNA
- inherited
two types- nuclear genome (found in nucleus), mitochondrial genome (found in mitochondria)
both useful for HID
what is the Human DNA
- most cells have 23 pairs of chromosome in the nucleus
chromosome composed of deoxyribonucleic acid
what is nuclear genome
- linear dna
- approx 3 billion base pairs long
- single cope per cell
what is mitochondrial genome
- circular dna
- 16, 569 bp long
what is diploid cells
- most cells are diploid
- two copies of each= 46 chromosomes
- somatic cells
what is haploid cells
- one copy of each = 23 chromosomes
- generally gamete cells- sperm and egg
- the basis of differences between individuals
- human DNA
DNA- consists of two long polymers running in opposite directions
- held together by hydrogen bonds
- coiled to form a double helix
- generally the most stable form
- each long polymer consists of sub units called nucleotides
- these are themselves made up of several constituent parts
What does a nucleotide consist of
- phosphate group
- 5 carbon sugar- deoxyribose
- one of nitrogen contains bases (A and T- G and C)
Genes
- human genome consists of over 25,000 genes
- up to 98% of human genome is non-coding (junk DNA)
Non- coding regions of DNA
sequence variation- sequences of bases present varies from individual to individual, less commonly used for genetic profiling
- length variation- within the junk DNA are areas where certain patterns are repeated, the number of repeats differ from individual o individual
DNA profiling- history
- first commercially used in 1987
- RFLP (restriction fragment length polymorphism)
- SLP and MLP (single and multi locus probe)
issues - these were time consuming
- require large amounts of target DNA
- not database able
- 1-6 weeks TRT
this is why PCR was invented
PCR
-a technique used to amplify DNA advantages - rapid - simple required only small amounts of DNA disadvantages - poor discriminating power - paved the way for current systems
Present DNA profiling
- focuses on variable repeating patterns (short tandem repeats)
- a form of mini satellite repeat commonly 2-6 bp in length
- technology counts the number of these repeats at several different areas or loci
the more loci we look at the better discriminatory power
SGM+
- current system used is the DNA 17
- Next generation multiplex- A 17- plex
- looks at 16 different STR areas within the genome all at the same time
- also looks at sex
- known as mulitplexing
- fully database able and compatible with the now updated UK database.
What is extraction and isolation?
once the body fluid has been found the cellular material must be extracted, it may be swabbed or the material cut and placed in a tube. The DNA must then be extracted from the cells and isolate to separate from other components and remove PCR inhibitors.
Basic process of extraction and isolation
- cell lysis- break open the cell
- removal of waste- removes inhibitors and dances
- purification/ concentration of DNA
quantification
once our DNA has been extracted we need to know how much we have
too much dna may result in overamplification
- dna profile becomes messy and difficult to interpret
- fails quality checks
too little dna
- low profile
- loss of information
picogreen
- a simple green dye that bind to DNA and fluoresces
- fluorescence measured and compared to a known standard
- not human specific and relatively inaccurate
PCR
denaturation- 95 degrees, double stranded dna melts
annealing- sample cooled to 66 degrees, allows primers to bind to specific areas
extension- sample heated to 72 degrees, enzyme copies DNA
process repeated numerous times
Post PCR
- each amplicon is labelled with a fluorescent tag
- enables us to visualise them later on
- however the fragments of DNA need to be separated using electrophoresis, this relies on the fact DNA is a charged molecule
electrophoresis
- charged particle migrates towards an electrode when placed in an electrical field
- DNA moves towards the anode
- larger fragments move more slowly and smaller fragments move more quickly
- fragments of the same size form discrete bands
electrophoresis media
agarose- a bit like clear jelly
polyacrylamide- much more effective as medium and most commonly used
POP- performance optimised polymer
