klausur

• variable number tandem repeats

• minisatellites

• larger repeat units

• up to 20kbp overall length

• short tandem repeats

• micro satellites

• shorter repeat units

• 100-400 bp overall length

• modern DNA profiling

• multilocus probes

• homologous to core sequence present in the repeat units of several VNTRs

• detect fragments of several VNTR loci at once

• give "bar code” -like pattern

• “genetic fingerprint”

• hybridize outside core region

• detect specific minisatellites

• only two bands per VNTR (= DNA profile)

a DNA fragment of a size characterized by an upper bound and a lower bound (electrophoretic ally non-resolvable size differences)

• alpha-Amylase

• Fluorescence

• Acid-phosphatase (AP)

• semenogelin (Sg)

• Fluorescence

• hemoglobin

• Fluorescence

• random breaks and deletions of DNA strand

• profile of locus falsely interpreted as homozygous

copy number =

number of starting molecules • 2^(number of PCR cycles)

• wear gloves and face mask (or pipette in safety cabinet)

• never re-use pipetten tips

• pipette slowly

• avoid touching inside of tube caps

• use different spatial areas for pre-PCR steps and for analyzing PCR products

16 (+AMELX/AMELY)

4.4•10^31 combinations

Gauß formula

n•(n+1)/2

additional 6bp in intron 1 at male gene (AMELY)

• cell lysis

• preservation of DNA integrity

• separation of DNA from other molecules

• heat

• acids

• DNases

• cell and tissue disruption

• removal of proteins and cytoplasmic contaminants

• concentration of DNA, removal of salts

• phenol method (outdated)

• chelex

• silica adsorption

  • columns

  • magnetic beads

• UV photometry

• fluorometry

• quantitative PCR (qPCR, real-time PCR)

• SYBR green -> simple and cheap

• TaqMan -> more specific

• agarose gel

• polacrylamide (PAA) gel

• denaturing PAA gel

• several DNA fragments amplified in a one-tube reaction

• standard technique for forensic DNA typing

• modern kits amplify up to 27 DNA loci

• avoid mia priming and primer dimerization

• start DNA synthesis (elongation) at elevated temperature

• contains PCR fragments of common alleles of a particular STR locus

• each locus has its own ladder

• has to be generated with same primers used in STR analysis

• runs separately

• typical for replication of short tandem repeats

• involves denaturing and re-annealing of repeat units

• the shorter the repeat unit, the less hydrogen bonds and the more likely stuttering is

D22S1045

allele drop-outs

• a true allele of one STR locus remains undetected

• a heterozygous locus appears homozygous

• too little amounts of DNA

• mutation in primer binding sites

• partial DNA degradation

• stuttering (+ADOs)

• DNA degradation

• mixtures

“whole genome amplification”

• natural genomic DNA is methylated, PCR-amplifies DNA is not

• a group of individuals of a single species

• live in same geographic area

• in which any pair of members can breed together

a group of people sharing common ancestry (e.g. east asian)

(copies/all alleles)

(individuals/all individuals)

• stable frequencies of alleles and genotypes

• a non-evolving population

• very large population

• random matings

• no migration

• no mutations

• no selective pressure

p^2 + 2pq + q^2 = 1

with p+q=1

• maternally inherited

• haplotype -> less variability than nuclear DNA

• mitochondrial genome is less vulnerable to degradation

• small amplifies (D loop) are tested

• present in higher copy number

• hair shafts and bone often contain intact mtDNA but often only degraded nuclear DNA

• buccal swabs

• blood

• teeth

• hair

• bone

1) isolation of DNA

2) PCR amplification of either HV1 or HV2

3) sequencing of HV1/2 -> PCR products

• profile present in patrilineage (no combination of two Y chromosome alleles)

• profiles are haplotypes -> RMP only by counting method

• much lower mutation rate than STRs (10^-8 vs 10^-3)

• shorter -> more reliably analyzed from degraded samples

• more than STRs distributed over genome (1 in every 500bp vs 1 in every 15000bp)

• can reveal more than just the identity of individuals

• identity-testing SNPs

• lineage-informative SNPs

• ancestry-informative SNPs

• phenotype-informative SNPs

• sanger sequencing

• single base extension (SBE; “mini sequencing”)

• next generation sequencing (NGS)

+

• highly informative

• better suited than STRs for typing degraded samples

-

• standardized assays not yet agreed upon

• standardized loci not yet agreed upon

• difficulty if develop >50 loci -multiplexes

• difficult to interpret mixtures

• forensic databases so far contain STR data

a group of organisms capable of interbreeding and producing fertile offspring

• based on morphology

• protein-based

• DNA-based

• suitable STR loci (many alleles, high heyerozygocity, robust PCR amplification possible, multiplexing of PCR possible)

• population data (allele frequencies for these STRs)