DNA Profiling

DNA ProfilingDNA Profiling

DNA ProfilingDNA Profiling

There are two methods used to carry There are two methods used to carry out DNA profilingout DNA profiling DNA profiling using a probeDNA profiling using a probe DNA profiling using PCRDNA profiling using PCR

Knowledge of this is not requiredKnowledge of this is not required It is the more common method now daysIt is the more common method now days

DNA profiling using a probeDNA profiling using a probe

This procedure relies on Minisatellites or This procedure relies on Minisatellites or Variable Number Tandem Repeats Variable Number Tandem Repeats (VNTR)(VNTR)

VNTR are sequences of DNA 10 – 100bp VNTR are sequences of DNA 10 – 100bp long that repeatlong that repeat

The number of repeats is inherited The number of repeats is inherited resulting in sections of DNA of different resulting in sections of DNA of different length (can be between 0.5kbp – 40kbp)length (can be between 0.5kbp – 40kbp)

DNA profiling using probes- DNA profiling using probes- procedureprocedure

Isolate DNAIsolate DNA DNA is isolated from sample using DNA is isolated from sample using

organic solvent which dissolves proteins organic solvent which dissolves proteins leaving DNAleaving DNA

If DNA quantity is insufficient PCR will be If DNA quantity is insufficient PCR will be performed (note this is different to the performed (note this is different to the method of DNA profiling using PCR)method of DNA profiling using PCR)

DNA profiling using probes – DNA profiling using probes – procedure cont…procedure cont…

Restriction enzyme digestionRestriction enzyme digestion Restriction enzyme which doesn’t cut in Restriction enzyme which doesn’t cut in

the minisatellite region is chosenthe minisatellite region is chosen Creates DNA fragment of different sizesCreates DNA fragment of different sizes


Gel electrophoresisGel electrophoresis Used to separate the different sized Used to separate the different sized

fragmentsfragments


Blotting DNA onto a filter (Southern Blotting DNA onto a filter (Southern Blotting)Blotting) DNA is denatured using, for example, a DNA is denatured using, for example, a

strong alkali – this creates single strong alkali – this creates single stranded DNAstranded DNA

Single stranded DNA (still in the Single stranded DNA (still in the electrophoresis pattern) is transferred to electrophoresis pattern) is transferred to a filtera filter



Hybridisation with probeHybridisation with probe Labelled (either radioactively or Labelled (either radioactively or

fluorescent) probe which binds to a specific fluorescent) probe which binds to a specific base sequence in the target fragments.base sequence in the target fragments.

Multi-locus probes bind at more than one Multi-locus probes bind at more than one site. Used where certainty needs to be site. Used where certainty needs to be very high.very high.

Single locus probes – bind to 1 sequence Single locus probes – bind to 1 sequence (giving 2 bands for each sequence)(giving 2 bands for each sequence)


Create autoradiographCreate autoradiograph Creates permanent record of the filter Creates permanent record of the filter

(usually using x-ray film)(usually using x-ray film) If using multi-locus, the filter is then If using multi-locus, the filter is then

washed and another probe is used to washed and another probe is used to add to the autoradiographadd to the autoradiograph

Interpreting Autoradiograph – Interpreting Autoradiograph – to establish paternityto establish paternity

After using a single locus probe 2 bands After using a single locus probe 2 bands are presentare present

As the VTNR forming the ‘band’ is As the VTNR forming the ‘band’ is inherited, 1 band is from maternal DNA, inherited, 1 band is from maternal DNA, the other from paternal DNA.the other from paternal DNA.

Therefore, a person that has the same Therefore, a person that has the same band as another person is potentially band as another person is potentially related.related.

A child’s bands should match one of the A child’s bands should match one of the mothers and one of the fathers.mothers and one of the fathers.

If this is not the case…….If this is not the case…….

Learning ActivitiesLearning Activities

Monograph pg 84 – 90Monograph pg 84 – 90 DART pg 87 – 88DART pg 87 – 88 Scholar 9.4Scholar 9.4 DNA profiling using Probes worksheetDNA profiling using Probes worksheet DNA profiling using PCR worksheet DNA profiling using PCR worksheet

(extension)(extension) Assignment questions 2001 Q11, 2002 Assignment questions 2001 Q11, 2002

Section C 1B, 2003 Q12, 2006 Q12Section C 1B, 2003 Q12, 2006 Q12

Documents

DNA Profiling