DNA Lecture 4 Flashcards
Human chromosomes display a one base difference in every ________ bases, on average
Every individual has some differences
1000
Base differences may or may not result in difference in amino acid sequences of proteins because
Not all segments of DNA code for proteins
There is more than one codon for most amino acids
The DNA sequence changes (single base differences) between individuals can affect recognition sites for
Restriction enzymes
Restriction enzyme sites are randomly made and eliminated
If a specific base sequence is present, the restriction enzyme which recognizes it will cleave the DNA molecule at the appropriate site and generate fragments of a specific length. However, if a restriction site is absent due to a base difference, different DNA restriction fragment lengths are formed.
!
Base sequence differences between individuals in a population are called
polymorphism
The variation in DNA restriction fragments between individuals are known as
Restriction Fragment Length Polymorphisms (RFLP)
The analysis of individual differences in RFLPs
In theory, there is a unique DNA fingerprint for every individual, the ____________ fragment pattern for each individual is unique
DNA finger printing
Restriction Enzyme
Detection of RFLPs relies on a specialized hybridization technique called
Southern Blotting
In DNA fingerprinting, first the DNA must be
Then the DNA must be
Then the DNA fragments must be
Then the DNA must be ______ so that it is single stranded
isolated from the rest of the cellular material in the nucleus
cut into several fragments of different sizes with restriction enzymes
Sorted by size
Denatured by soaking the agarose gel in alkaline solution
After the DNA is denatured in fingerprinting, the DNA is blotted onto a nylon membrane. The strands of DNA bind to the positively charged groups in the nylon membrane because of..
The negative charge on the DNA bases
After blotting the DNA, the membrane is immersed in a solution of radiolabeled DNA probe.
The probe is a 32P labeled oligonucleotide whose sequence is complementary to a segment of known sequence of DNA.
The oligonucleotide with hybridize to the fragment that contains its
Complimentary Sequence
Once the probe is bound to the DNA in DNA fingerprinting, the result is washed and analyzed via autoradiography.
The fragment length is determined from the position on the gel compared to DNA markers of
Known Length
Usually DNA fragments resulting from DNA fingerprinting (aka restriction digest) for paternity testing are separately analyzed with several 32P labeled oligonucleotide probes. Combining the use of these makes the test extremely selective.
Each gene fragment analyzed has a different paternity index, from those a combined paternity index is reported. A probability is calculated
Many mutations either create a new restriction site in a genome or destroy a previously existing site. If a mutation occurs within or near a particular gene, the restriction fragmentation pattern will change.
This change can be determined by ______ with a sequence that lies in the same region of the genome
Southern blotting
This disease has an altered restriction pattern when DNA is probed for part of the gene for the beta-globin subunit, enabling prenatal diagnosis
Sickle cell disease
Other ex:
Cystic Fibrosis, Huntington’s, cancer, manic depression
In a classic example of RFLPs, the semen sample obtained from a rape and murder victim was compared with samples from the victim and two suspects. The samples were cleaved by ________ and separated by ___________ and processed by ________
The samples were cleaved by endonucleases and separated by gel electrophoresis and processed by the Southern blotting procedure
Name the two disadvantages of the Southern blotting method in a crime scene
Southern blot requires fresh DNA samples and larger amount of DNA than those usually found on a crime scene
The need for more sensitive DNA typing methods led to a focus on the _________ and on ______
polymerase chain reaction (PCR) and on STRs
Most genetic fingerprinting done today analyzes regions of our genome referred to as _________ that have highly variable repeating sequences
Minisatellites
The minisatellites have _______ most commonly 4 bases long that are repeated several times, such as (TAGA)m
Short Tandem Repeats (STRs)
____ is observed between the number of repeats (n) in each STR region
Strs represent about ____% of all human DNA
Polymorphism
3%
By analyzing the number of repeats in many minisatellites, a unique profile for each individual can be obtained.
_______ is readily applicable to STR analysis
Polymerase Chain Reaction (PCR)
This procedure makes it possible to isolate and analyze the DNA in a very small tissue sample (e.g. one human hair, a drop of blood, a small semen sample, or samples that are many years old)
With the increased sensitivity, forensic scientists focused on STRs rather than RFLPs
PCR
In this first step of PCR, DNA strands are separated by
In the second step, strands are annealed to an excess of short DNA _____ that flank the region to be amplified
New DNA is synthesize by ____
heating
Primers
Polymerization
