DNA Fingerprinting Flashcards
why is DNA unique in individuals
as the genome in a human has a specific order of nucleotides and they have a unique pattern of polymorphic sequences in his/her chromosomes (exception: twins)
what are restriction enzymes +function +example
- they are molecular scissors (enzymes), made by bacteria, that cut at a restriction site in a nucleotide sequences
example: EcoRl
what is DNA’s charge + reason
-negative
how and why can DNA fragments move in an electrical field
- due to the negative CHARGE of the DNA, and because opposites attract, there is a positive end at the opposite of the electrical field that pulls the DNA fragments
- the smaller the piece, the further it moves down the electrophoresis gel
why do we need to use the same restriction enzymes for a crime scene investigation
- to recognize restriction sites + cut
- the site where they cut generate specifically sized fragments of DNA that are unique to an individual
what are polymorphic DNA sequences +effect on restriction enzymes
- regions on human chromosomes that exhibit a great deal of diversity
- they can change the size of the restriction enzymes
what are short tandem repeats (satellite DNA)
- they are polymorphic sequences that lie close to each other on the chromosome that repeat for a while
- highly variable between individuals
where are STR’s found
in the inter-genie region
what are STR’s used for
- forensic identification
- paternity testing
- genetic disease diagnosis
how can STR’s be used
- to establish human identity (fingerprinting)
what is a restriction site
- a specific sequences of base pairs in DNA
- its a palindromic sequence (read the same way right to left as it is done left to right)Ex
what is the restriction site for the enzyme EcoRL
AATT and TTAA
how do restriction enzymes cut
- if it cuts at one location, two fragments produced
- if it cuts at two locations, three fragments produced
what happens when two pieces of DNA are identical
-a specific restriction enzymes will cleave the two DNA samples in the same way, generating two fragments of the same size
what are the steps of DNA profiling
- DNA is amplified by PCR reaction (polymerase chain reaction)
- DNA is cut up into short fragments using restriction enzymes
- Gel Electrophoresis is used to separate fragments by size and charge
what do you do during the PCR reaction
- Denaturation: temperature raised to separate the DNA strands at 90.C
- Annealing: primers allowed to anneal as temperature drops (50.C)
- Extension: Tagpolymerase elongates new strands of DNA
process of gel electrophoresis lab
- In a gel electrophoresis lab a mixture of restriction enzymes, DNA samples and loading dye (to indicate where the fragments are and to weigh down the mixture) are inserted into their respective wells in an agarose gel (this mixture is measured with a micropipette which inserts indicated amounts into microcentrifuge tubes, the mixture is then mixed and collected to the bottom of the microcentrifuge tube using a pulse centrifuge).
- When the restriction enzymes are inserted into the tubes and once they are all in the microcentrifuge tube rack, they are to be incubated for 45 minutes at 37°C in a water bath so that the enzymes can work under an equal and optimum reaction rate.
why is a buffer used in the lab (when gel is submerged in buffer solution)
-optimal PH for lab for the enzymes (stabilize experiment)
why do we use a standard solution
- A standard solution, the DNA from the crime scene and the DNA samples of crime suspects are inserted into these wells.
- the standard solution is used for comparative reasons. Once the lab is over and the bands of the DNA solutions appear the standard solution shows bands where the number of base pairs is already known.
why water bath
Water bath and consistent temperature regulation used to make the rate of reaction of the restriction enzymes consistent and equal amongst different samples, and use a temperature which works best for the restriction enzyme.
