Forensic genetics is part of forensic medicine Identification

Forensic genetics is part of forensic medicine Identification of • crime perpetrator. • Paternity • Unidentified bodies First system used: ABO blood groups (Landsteiner, 1900), with other blood groups (MN, Rh) and other proteins.

Blood groups Phenotype Genotype 0 A B AB i/i IA/IA o IA/i IB/IB o IB/i IA/IB 3 allelic variants: IA; IB (dominant); i (recessive)

Genitori Figli Possibili entrambi A A, ZERO entrambi B B, ZERO entrambi AB A, B, AB entrambi ZERO uno A l'altro B A, B, AB, ZERO uno A l'altro AB A, B, AB uno A l'altro ZERO A, ZERO uno B l'altro AB A, B, AB uno B l'altro ZERO B, ZERO uno AB l'altro ZERO A, B

Phenotype M N MN Genotype LM /LM LN/LN LM/LN Two allelic variants: LM ; LN (codominant) Phenotype Rh+ Rh- Genotype D /D o D/d d/d Two allelic variants: D (Dominant); d D

DNA fingerprinting Pattern of bands specific for each individual

Advantage Discriminatory power PROBLEMS High amount of DNA required Time consuming and manual Samples with degraded DNA Statistics (probatory value)

1990 PCR in forensic medicine Little DNA required. Use with microsatellites: high discriminatory power.

The Organization of Human Genome: Minisatellite DNA 2 major family Medium sized arrays of tandemly repeated DNA, up to 20 kb Basic sequences are usually very simple Basic block of 6 bp (telomere DNA), 64 bp (hypervariable) Usually not transcribed Hyper variable family: more than 1000 different array in the genome Highly polymorphic, in composition (restriction sites) and in length: DNA fingerprinting, Using a single DNA probe containing the common core sequences Found near telomeres but also in chromosomal locations

The Organization of Human Genome: Microsatellite DNA (Short Tandem Repeat) small arrays of tandemly repeated DNA, about 150 bp Detected in the genome of every organism Highly instable and polymorphic In human: Scattered across different chromosomal locations Mononucleotide repeats, A and T runs are very common Dinucleotides repeats CT/AG are less common

Amplification of microsatellite fragment Primers in GREEN Amplified fragment in GREY Microsatellite in BLU

Highly heterozygous in the population (Discriminatory) Short PCR products (Efficiency) Separable on Gel or Capillary Electrophoresis Low mutations rate (Automation)

Multiplex PCR Capillary Electrophoresis Separation based on length of the fragments and labelling of the fluorophores

NAME OF STR; (Number of repeats) Optimal annealing temperature must be similar

Global. Filer Express

Promega Power. Plex Fusion

Power. Plex Fusion

Probability of Identity (Independent alleles) Pi(locus 1) x Pi(locus 2) Combined 6. 18 x 10 -27 3. 34 x 10 -24 3. 71 x 10 -26 3. 09 x 10 -26

Probability match 1 / 10 x 1 / 111 x 1 / 20 1 / 22, 200 1 / 100 x 1 / 14 x 1 / 81 1 / 113, 400 1 / 116 x 1 / 17 x 1 / 16 1 / 31, 552 1 / 79, 531, 528, 960, 000

Problem 1: stutter

Problem 2: Allele dropout (kit related)

Problem: samples Degraded DNA

Problem: samples Low template DNA (<100 pg) #1 #2 #3

Problem sample: Mixed samples

Sono stati sviluppati kit per la determinazione del profilo Y -specifico kit Yfiler della Life Technologies amplificazione di 17 loci STR kit Y-specifico della Promega si ottiene un profilo a 23 loci STR