113 Questions
How many hydrogen bonds are between adenine and thymine in DNA?
Two
Which bases are paired by three hydrogen bonds in DNA?
Cytosine and guanine
In the DNA double helix, which direction do the sugar-phosphate backbones run?
One in the 5' to 3' direction, the other in 3' to 5' direction
What enables the optimization of energy levels within the DNA double helix?
Complementary base-pairing of A with T and G with C
How often do twists or turns occur in the sugar-phosphate backbones of the DNA double helix?
Every ten base pairs
What provides a template for creating an exact copy of the DNA molecule during replication?
Each DNA strand in the double helix
Where is DNA packed into tightly coiled structures called chromatin?
In the cell's nucleus
What is the purpose of Platelia™ Aspergillus Ag test?
To detect galactomannan Aspergillus antigen
What proteins are separated in SDS-PAGE in the Western blot technique?
Antibodies from test serum
Which disease detection is NOT mentioned for serological tests in the text?
Influenza
What is the purpose of FISH (in-situ hybridization)?
To detect specific pathogens without isolating DNA or RNA
What is the method used to estimate the time of infection in Lyme boreliosis?
Antibody concentration and IgM to IgG ratio
Which method allows you to amplify a specific fragment of DNA by several thousands base pairs?
PCR - amplification
Which antibody is produced earlier in the course of Lyme boreliosis?
IgM
What is the purpose of reverse transcription in RT-PCR?
To transcribe RNA into DNA
What infectious agent is NOT included in the serological methods mentioned?
E. coli
Which technique is used for phylogenetic studies in mycology?
PCR - polymerase chain
What type of infection is detected using the Platelia™ Aspergillus Ag test?
Fungal infection
What is the key feature of PCR that allows for the amplification of DNA?
Repeated heating of the sample
'What is the main advantage of serological tests mentioned in the text?'
Quick diagnosis of many infectious diseases
Which method is used for the detection of Cryptosporidium, Giardia, and Microsporidium in environmental studies?
RT-PCR - reverse transcription
What does RT-PCR stand for?
Reverse Transcription Polymerase Chain Reaction
Which technique allows the transcription of RNA into DNA before amplification?
RT-PCR - reverse transcription
What is one advantage of serological methods for antigen detection?
Fast detection
What is a disadvantage of serological methods due to antigenic similarity between microorganisms?
Cross reactions
In molecular methods used in microbiological diagnostics, what element is analyzed?
DNA sequences
What type of diagnostics are molecular methods commonly used in besides microbiology?
Genetic diagnosis of diseases
What is the main focus of molecular methods in parasitological diagnostics?
Analysis of DNA sequences specific to pathogens
Why might serological methods pose a challenge in cases of lack of antibodies?
False positive results
What is a common issue with serological methods related to high sensitivity?
Low specificity
How are molecular probes described in the context of molecular methods?
DNA sequences characteristic of the species sought
What is a disadvantage of DNA molecule analysis mentioned in the text?
Need for sterile conditions
Why is there a risk of contamination in DNA molecule analysis?
The immediate vicinity of the laboratory
What is a challenge of gene amplification mentioned in the text?
Need for appropriate primers
What is the main advantage of DNA molecule analysis according to the text?
High yield of gene copies
Why is knowledge of the gene sequence important in DNA molecule analysis?
To select appropriate primers
What additional analyses are needed to visualize the effects of DNA molecule analysis?
Gel separation and staining
Which aspect makes DNA molecule analysis time-consuming?
'Rate' of test gene amplification time of 2-3 hours
What is the purpose of the annealing step in the PCR process?
To attach the primer sequences to the DNA strand
During which step of PCR is the sample heated to 95°C to melt the double helix?
Denaturation
What determines the temperature used in the annealing step of PCR?
The content of GC pairs
What is the main advantage of controlling the PCR amplification process?
Ensuring exponential amplification
What is a potential disadvantage of PCR mentioned in the text?
Amplifies non-specific products
What is the purpose of using multiple primer pairs in Multiplex PCR?
To allow for the amplification of two different genes simultaneously
Which statement best describes the process of annealing in PCR?
It is the stage where primers bind to the complementary sequences on DNA strands.
What contributes to the increased sensitivity and specificity in Nested PCR?
Amplifying the sequence first with 'external' primers and then 'internal' primers
In PCR, what is the purpose of temperature control during the amplification process?
To ensure optimal binding of primers to DNA strands
What is a key advantage of using nested PCR in diagnostics compared to traditional PCR?
Increased sensitivity and specificity
What is the purpose of the annealing step in the PCR process?
Allowing primers to bind to the complementary sequences of DNA
Which of the following is a disadvantage of PCR mentioned in the text?
High cost of materials
What is the role of the extension/elongation step in PCR?
Synthesizing new DNA strands from primers
Which factor contributes to the frequent false-negative results in PCR?
Lack of standardization
What is one advantage of PCR mentioned in the text?
Lack of material needed for testing
What is a common disadvantage of the PCR technique mentioned in the text?
Low yield of tested gene copies
What aspect of DNA molecule analysis requires the need for sterile conditions?
Annealing with appropriate primers
Which factor contributes significantly to the complexity of implementing DNA molecule analysis?
Selectivity
What is a critical requirement for successful gene amplification using PCR?
High temperature control during annealing
Which feature of PCR contributes to the possible contamination risks during DNA molecule analysis?
The need for selecting appropriate primers
What is the purpose of the annealing step in the PCR process?
To lower the temperature for primers to bind to the DNA template
Which factor is crucially controlled during the extension/elongation phase of PCR?
Maintaining a stable temperature for DNA synthesis
What is a primary advantage of PCR in comparison to traditional DNA replication methods?
Allows for rapid and specific amplification of DNA fragments
What is a potential disadvantage associated with poor temperature control in the PCR process?
Overheating and denaturation of DNA polymerase
In PCR, what role does the polymerase chain reaction play during the amplification process?
Amplifies specific DNA regions through repeated cycles
What is the purpose of the annealing step in PCR?
To allow bases to pair by three hydrogen bonds
Why is controlling the PCR amplification process advantageous?
To prevent non-specific amplification and ensure specificity
What is a potential disadvantage of DNA molecule analysis mentioned in the text?
Risk of contamination
Which of the following is NOT a common issue related to high sensitivity in serological methods?
Lack of antibodies in immunocompromised individuals
What aspect of PCR allows for the amplification of DNA sequences?
Amplification process control
What is the purpose of the annealing step in the PCR process?
To allow primers to bind to the template DNA
During PCR, what is the role of DNA polymerase in the extension/elongation step?
Synthesizing a complementary DNA strand
What determines the temperature used in the annealing step of PCR?
The melting temperature of primer-template DNA duplex
What is one advantage of controlling the PCR amplification process?
Avoiding nonspecific amplification
What is a potential disadvantage of PCR as mentioned in the text?
Risk of contamination
What provides a template for creating an exact copy of the DNA molecule during replication?
Template DNA
Why might serological methods pose a challenge in cases of lack of antibodies?
They rely on detecting specific antigens
What is a common disadvantage of PCR mentioned in the text?
Frequent false negative results
Which factor plays a crucial role in controlling the speed of obtaining PCR results?
The intensity of fluorescence growth
Why does poor temperature control pose a significant challenge in the PCR process?
It affects the amplification efficiency
What advantage does PCR offer in terms of material needed for testing?
Small amount of material needed for testing
Which aspect is an advantage of PCR when considering the possibility of using different materials?
Speed of obtaining the result
What is the purpose of the denaturation step in the PCR process?
To melt the double helix and break hydrogen bonds
During the annealing step in PCR, what does the temperature depend on?
The length of the primers and the GC pairs content
In PCR, what happens during the extension/elongation step?
Synthesis of a complementary strand by a polymerase
What is controlled during the amplification process in PCR?
The number of repetition cycles
What is a primary advantage of PCR mentioned in the text?
Exponential amplification of products
What is a key disadvantage of PCR mentioned in the text?
Risk of contamination
Why is the selectivity of PCR considered a disadvantage?
It allows the amplification of only the target sequence
What is the role of temperature control during the annealing step of PCR?
To differentiate between bacterial contaminants
How does control over the extension/elongation phase benefit PCR?
Ensures accurate replication of DNA sequences
Which of the following is NOT an advantage of PCR as mentioned in the text?
Ability to work in non-sterile conditions
What is the purpose of the annealing step in PCR?
To attach primers to the DNA template
In PCR, what is the role of DNA polymerase in the extension/elongation step?
Adding nucleotides to the growing DNA strand
What enables the optimization of energy levels within the DNA double helix during PCR?
Maintaining the annealing temperature
Which factor contributes to the increased sensitivity and specificity of nested PCR?
Employing external primers in both reactions
What is a potential advantage of PCR mentioned in the text?
Amplification of multiple genes simultaneously
What is the purpose of the extension/elongation step in PCR?
Enabling the DNA polymerase to synthesize new DNA strands from primers
What determines the temperature used in the annealing step of PCR?
The sequence of the primer binding sites on the DNA template
Why is controlling the PCR amplification process advantageous?
To ensure accurate and reproducible amplification of DNA fragments
What is a potential disadvantage of using PCR as mentioned in the text?
High risk of contamination during amplification
What is one advantage of controlling the amplification process in PCR?
Ensuring uniform distribution of DNA fragments
What is a critical factor controlled during the extension/elongation phase of PCR?
Temperature
Which factor contributes significantly to the complexity of implementing DNA molecule analysis?
DNA polymerase choice
What is a common disadvantage of the PCR technique mentioned in the text?
Risk of contamination
Why is controlling the PCR amplification process advantageous?
To ensure specificity
What contributes to the increased risk of contamination during DNA molecule analysis?
Polymerase chain reaction
During PCR, what is the main purpose of the annealing step?
Allowing primers to bind to complementary sequences
In PCR, what is the role of DNA polymerase during the extension/elongation step?
Synthesizing a complementary DNA strand
What is a critical factor that poor temperature control poses during the PCR process?
Reduced enzyme activity
Which factor plays a crucial role in controlling the speed of obtaining PCR results?
The duration of the denaturation step
What is an advantage of controlling the amplification process during PCR?
Reduced non-specific amplification
What is a common disadvantage associated with PCR amplification?
Potential contamination risks
Why is temperature control crucial in the PCR process?
To maintain enzyme activity
What is a key advantage of using PCR compared to traditional DNA replication methods?
Rapid and target-specific amplification
Study Notes
Here are the study notes in detailed bullet points:
DNA Structure
- Adenine (A) and Thymine (T) are paired by two hydrogen bonds, while Cytosine (C) and Guanine (G) are paired by three hydrogen bonds (complementary base pairing)
- The two DNA strands run in opposite directions (antiparallel) to help the bases in each base pair fit into the double helix
- The width of each base pair remains the same, keeping a consistent distance between the sugar-phosphate backbones along the length of the DNA molecule
- Twists or turns in the sugar-phosphate backbones of the double helix occur every 10 base pairs, maximizing the efficiency of base-pair packing
DNA Replication
- Each strand of DNA serves as a template for creating an exact or identical copy of the DNA molecule
- Separation of the two strands allows for duplication of the DNA molecule
Molecular Diagnostics
- Molecular methods are used in microbiological diagnostics (virology, bacteriology, mycology) and parasitological diagnostics of humans and animals
- Molecular methods are also used in the genetic diagnosis of hereditary diseases and human cancers
- The element analyzed in molecular methods is the presence of DNA sequences (or RNA in some methods) in the tested material characteristic of the sought species of the microorganism suspected of causing infection
PCR (Polymerase Chain Reaction)
- A laboratory technique that amplifies a specific DNA sequence
- Consists of three stages: denaturation, annealing, and extension/elongation
- PCR is used in diagnostics of various diseases and infections, such as Candida, Aspergillus, and Cryptococcus
- Real-time PCR observes the amplification in real-time using fluorescent probes or markers
Types of PCR
-
RT-PCR (Reverse Transcription Polymerase Chain Reaction): used for RNA viruses and for detecting genetic material in the environment
-
Nested PCR: uses two different primer pairs to increase sensitivity and specificity of the reaction
-
Multiplex PCR: allows for the multiplication of more than one sequence in a single reaction### PCR (Polymerase Chain Reaction)
-
A laboratory technique that mimics the process of DNA replication in vitro
-
Allows amplification of specific DNA fragments (tens to thousands of base pairs)
-
Involves DNA isolation, PCR amplification, and product distribution and visualization on agarose gel
Advantages of PCR
- High sensitivity
- High yield (10^6 - 10^9 copies of the tested gene)
- Rapid test (2-3 hours)
- Simple implementation
- Selectable primers for the reproduced gene
Disadvantages of PCR
- Risk of contamination (e.g., with the genetic material of the person performing the analysis)
- Selectivity (only the selected sequence is reproduced)
- Need to work in sterile conditions
- Need to select appropriate primers for the reproduced gene
- Need to perform additional analyses to visualize the obtained effects (gel separation, restriction enzyme digestion, staining)
FISH (Fluorescence In-Situ Hybridization)
- A technique used for the detection of specific DNA sequences
- Involves hybridization of nucleic acids to probes
- Applications:
- Detection of Candida, Cryptococcus, Histoplasma, and other microorganisms
- Detection of invasive infections induced by Plasmodium, Leishmania, Trypanosoma in the blood and cerebrospinal fluid
- Detection of Cryptosporidium, Giardia, Microsporidium
RT-PCR (Reverse Transcription Polymerase Chain Reaction)
- A variant of PCR that involves the reverse transcription of RNA into DNA
- Used for the detection of RNA viruses and the analysis of gene expression
Molecular Methods
- Used in microbiological diagnostics (virology, bacteriology, mycology) and parasitological diagnostics of humans and animals
- Also used in the genetic diagnosis of hereditary diseases and human cancers
- Involve the evaluation of the presence of DNA sequences (or RNA) in the tested material characteristic of the sought species of the microorganism suspected of causing infection
Learn about the advantages and disadvantages of using serological methods for antigen detection and molecular methods for microbiological diagnostics. Understand the benefits and limitations of each approach in identifying infections and antibodies.
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