Genetics - Genetic Testing Flashcards
What is karyotyping?
It is a technique that produces an image of an individual’s chromosomes
The chromosomes are ordered in size and grouped together in their pairs
What does karyotyping detect?
It is used to detect abnormal numbers or structures of chromosomes
What are the three types of chromosome structures?
Metacentric
Sub-metacentric
Acrocentric
What are metacentric chromosomes?
Those which have the centromere in the centre so that both sections are of equal length
What are submetacentric chromosomes?
Those which have the centromere slightly offset from the centre leading to a slight asymmetry in the length of the two sections
What are acrocentric chromosomes?
Those which have a centromere which is severely offset from the centre leading to one very long and one very short section
What are FISH studies?
It is a technique that identifies where a particular gene falls within an individual’s chromosomes
How do we conduct FISH studies?
We use fluorescent probes that are complementary to the genes we are trying to locate within the chromosome
What are the four types of FISH probes?
Unique Sequence Probes
Centromeric Probes
Telomeric Probes
Whole Chromosome Probes
What are unique sequence probes?
These probes bind to specific small segments of certain chromosomes
What two chromosomal abnormalities are detected by unique sequence probes?
Microdeletion
Microduplication
What are centromeric probes?
These probes target the centromeric region of the particular chromosome
What is the function of centromeric probes?
They allow us to enumerate the number of centromeres and thus chromosomes of a particular kind
What are telomeric probes?
These probes bind to telomeres, which are the repeat sequences found at the end of the chromosomes
What are whole chromosome probes?
These probes are usually collections of smaller probes, each of which bind to a different sequence along the length of a given chromosome
What two chromosomal abnormalities are detected by whole chromosome probes?
Translocations
Rearrangements
What does PCR testing detect?
It is used to detect where DNA is replicated, allowing genes of interest to be screened for
What are the five substances required to carry out PCR?
Sequence Information
Oligonucleotide Primers
DNA
Nucleotides
DNA Polymerase
What are the three PCR steps?
Denaturation
Annealing
Elongation
What is denaturation?
This is where the sample is heated to allow the double-stranded DNA to be separated into single strands
This is due to the fact that the heat is able to break the hydrogen bonds between the nucleotide base pairs in the two strands of DNA
What temperature is used during denaturation?
93-95 degrees
What is annealing?
This is where the sample is cooled
The DNA primer will bind to the target DNA sequences on the two DNA strands
What temperature is used during annealing?
50-70 degrees
What is elongation?
New strands of DNA are made using the original strands as templates
A DNA polymerase enzyme joins free DNA nucleotides together
The order in which the free nucleotides are added is determined by the sequence of nucleotides in the original DNA strand
What temperature is used during elongation?
70 - 75 degrees
What is the result of one PCR cycle?
Two double-stranded sequences of target DNA, each containing one newly made strand and one original strand
What is gel electrophoresis?
It is used to separate DNA, RNA fragments or proteins based on their size and charge
How do we conduct gel electrophoresis?
It involves running a current through an agarose gel containing the molecules of interest
Based upon their size and charge, the molecules will travel through the gel in different directions and speeds, allowing them to be separated from one another
What is restriction fragment length polymorphism (RFLP) analysis?
It is a technique that exploits variations in homologous DNA sequences, known as polymorphisms
How do we conduct RFLP analysis?
It involves a fragmenting a sample of DNA with the application of enzymes, known as restriction endonucleases
These enzymes can selectively cleave a DNA molecule wherever a short, specific sequences is recognised
We then transfer these DNA fragments to a blotting sheet, where we add a labelled DNA probe
This DNA probe determines the length of the fragments which are complementary to the probe
A restriction fragment length polymorphism is said to occur when the length of a detected fragment varies between individuals, indicating non-identical sequence homologies
What are restriction endonucleases?
They are enzymes from bacterial cells which are able to recognise specific DNA sequences and always cut DNA at the same site
What is an amplification refractory mutation system (ARMS)?
It is a method for detecting any mutation involving single base changes or small deletions
How do we conduct ARMS?
It involves the application of sequence-specific PCR primers that allow amplification of DNA when the target allele is contained within the sample
Therefore, the presence or absence of a PCR product allows us to determine the presence or absence of the target allele
If the target allele is present, then amplification of the DNA sequence will occur
What does ELISA testing detect?
It is used to screen for specific antibodies in a patients serum
What does Eastern blot testing detect?
The post-translational modification of proteins
What does Western blot testing detect?
It is used to separate and identify proteins
What disease is detected using Western blot testing?
Mad cow disease
