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What invention in 1983 by Kary Mullis revolutionized molecular biology research?
The polymerase chain reaction (PCR).
What does PCR stand for?
Polymerase Chain Reaction.
What are PCR products also known as?
Amplicons.
What are the three main stages of PCR?
Denaturation, annealing, and extension.
What are primers in a PCR reaction?
Short, single strands of DNA that match and bind to the target sequence.
What temperature is typically used for denaturing DNA in PCR?
94°C.
What is the function of Taq DNA polymerase in PCR?
It extends the primers and synthesizes new DNA strands.
How many cycles are typically run in a PCR reaction to amplify the target sequence?
Between 25 to 40 cycles.
What is the melting temperature (Tm) in PCR?
The temperature at which half of the DNA primers dissociate from the template.
What is the purpose of PCR optimization?
To determine the best combination of components and conditions for each specific PCR reaction.
What is Reverse Transcription PCR (RT-PCR) used for?
To convert mRNA into complementary DNA (cDNA) for subsequent PCR amplification.
What does qPCR stand for?
Quantitative Polymerase Chain Reaction.
What real-world application does PCR provide in forensics?
DNA profiling to identify suspects based on biological samples.
What technique is used to analyze PCR products after amplification?
Agarose gel electrophoresis.
Why is the Tm of two primers in PCR ideally similar?
To ensure only one annealing temperature can be used for the reaction.
What is multiplex PCR?
A PCR technique that amplifies multiple target sequences in the same reaction tube.
What is the role of magnesium chloride (MgCl2) in a PCR reaction?
It serves as a necessary cofactor for DNA polymerases to function.
How is PCR applied in agriculture?
To detect genetically modified organisms (GMOs) and to create genetically modified plants.
What is the significance of STR analysis in forensics?
It allows fast and highly discriminatory DNA profiling.
How does PCR contribute to medical diagnostics?
It enables rapid detection and diagnosis of diseases such as viral infections.
What is next-generation sequencing?
A method intended to sequence entire genomes faster and at a lower cost than traditional sequencing methods.
What is the purpose of a master mix in PCR?
To combine all necessary components of a PCR reaction to simplify multiple reactions.
What is the advantage of using real-time PCR over conventional PCR?
It allows quantification of DNA as PCR products are generated.
What biological mark do primers used in PCR require?
They must match specific sequences at each end of the target DNA region.
What is the expected outcome after 40 cycles of PCR amplification theoretically?
Billions of copies of the target DNA sequence.
What characteristic of DNA polymerases is essential for PCR?
Thermostability; they must withstand high temperatures during the denaturation step.
In what way can PCR be used in animal conservation?
To identify biological samples, such as ivory or bear gall bladders, to combat illegal wildlife trade.
What is an application of PCR in human migration studies?
To trace the history of mitochondrial DNA mutations.
What is the benefit of using agarose gels with higher percentages for PCR products?
To analyze smaller PCR products, often less than 1,000 base pairs.
How does PCR facilitate the production of biological drugs?
By cloning the genes of interest for therapeutic antibodies.
What does the term 'pharming' refer to?
The use of genetically modified animals to produce pharmaceutical products in their milk.
How do forensic DNA databases raise privacy concerns?
They contain DNA profiles of individuals, including those who have not been convicted of a crime.
What is the purpose of using negative controls in PCR reactions?
To ensure that there has been no contamination affecting the results.
What is an advantage of using a heated lid on thermal cyclers?
To prevent evaporation of samples during PCR reactions.
What is the main reason for the success of PCR in forensic science?
Its ability to amplify very small quantities of DNA.
What is the role of the Alu sequence in human genome studies?
To trace human migration and genetic variation.
What is a key characteristic of next-generation sequencing compared to traditional methods?
It is significantly faster and more cost-effective.
Why are aerosol-barrier pipet tips recommended in PCR?
To prevent contamination of PCR reagents.
How did PCR impact the Human Genome Project?
It provided the means to rapidly amplify and sequence DNA.
How is PCR used in paternity testing?
By comparing the number of STR repeats between a child and potential parents.
What temperature range is typically optimal for primers to anneal?
50-60°C.
What is the potential consequence of undetected contamination in PCR?
Amplification of incorrect DNA, leading to false results.
What is the significance of the Combined DNA Index System (CODIS) in forensic analysis?
It utilizes specific STR loci to create profiles for identification.
Which reaction component is NOT essential for PCR?
RNA primers; DNA primers are used instead.
What innovative method did thermal cyclers replace in PCR?
Manual transfers between separate temperature-controlled water baths.
What is a potential relevance of PCR in diagnosing viral outbreaks?
It allows for the quick confirmation of infections such as influenza.
How does the length of a PCR primer typically range?
17-28 base pairs.
What property of primers enhances the specificity of PCR?
The presence of a GC clamp at the 3' end.
What is the importance of dideoxynucleotides (ddNTPs) in cycle sequencing?
They terminate DNA strand elongation, allowing for the determination of DNA sequence.
What type of enzyme, aside from Taq polymerase, is often used in high fidelity PCR?
Pfu polymerase, which has proofreading abilities.
What is one potential limitation of using multiplex PCR?
It requires careful design of multiple primer sets to avoid non-specific amplification.
What role does PCR play in detecting GMOs in food?
It amplifies specific genetic modifications from food samples for identification.
Why is it crucial to optimize PCR reactions?
To achieve reliable amplification of the target DNA under specific conditions.
What challenge may arise when attempting to amplify degraded DNA?
The target sequences must be short to allow successful primer binding.
What is the aim of genetic modifications in crops?
To improve traits such as pest resistance and herbicide tolerance.
How can the number of loci tested in STR analysis affect discrimination power?
Increasing loci reduces the likelihood of random matches between DNA profiles.
What is the expected amplification factor after 35 cycles of PCR?
Approximately 4 billion times more copies of the target sequence.
What are the primary components of a standard PCR reaction?
Template DNA, nucleotides, primers, buffer, magnesium chloride, and DNA polymerase.
What was the goal of the Human Genome Project?
To map and sequence the entire human genome.
How do fluorescent dyes function in real-time PCR?
They bind to double-stranded DNA and emit fluorescence during amplification.
What is the relationship between alleles and genotypes?
An allele represents a specific variant at a genetic locus, while a genotype is the combination of alleles inherited.
What are the biological implications of PCR's ability to amplify DNA?
It enables detailed genetic analysis and applications in various fields such as medicine, forensics, and agriculture.
Why is quality control needed during PCR setup?
To verify that experimental conditions are met and contamination is avoided.
In what way can human migration studies benefit from PCR technology?
PCR can analyze genetic markers to trace lineage and ancestry.
What is the potential economic benefit of transgenic crops?
Increased agricultural yield and reduced costs of pest management.
What is the underlying technique in DNA microarray technology?
PCR amplifies DNA to produce sufficient material for analysis on microarrays.
What does the term 'biological fingerprinting' refer to?
The use of genetic analysis techniques to identify individuals based on their DNA.
How does PCR help in recognizing the presence of RNA viruses?
By using RT-PCR to convert RNA into DNA for subsequent amplification.
What potential application does PCR have in wildlife conservation?
Identifying illegal animal parts to aid in conservation efforts.
How does the presence of a GC clamp affect primer efficiency?
It enhances binding strength of the primer to the target DNA.
What novel approaches are being developed in genome sequencing arising from PCR advancements?
Techniques such as next-generation sequencing for faster and cheaper genomic analysis.
Why might PCR fail with too much or too little template DNA?
Excess DNA can inhibit reactions while insufficient DNA may not yield detectable products.
What is the purpose of using positive and negative controls in PCR assays?
To ensure amplification is successful and to check for contamination.
In what ways has PCR simplified the process of genetic testing for diseases?
By allowing rapid and specific amplification of genetic sequences related to diseases.
What is one drawback of using conventional PCR in genetic screenings?
It provides only qualitative results rather than quantification of DNA.
What is the importance of dNTPs in PCR reactions?
They serve as building blocks for new DNA strands during amplification.
What is the difference between conventional PCR and RT-PCR?
RT-PCR converts RNA into DNA before amplification, while conventional PCR amplifies DNA directly.
What is a key factor determining the success of a PCR reaction?
The specificity and design of primers used in the reaction.
How does PCR contribute to the field of personalized medicine?
It allows for tailoring medical treatments based on individual genetic profiles.
What essential factor must be considered when designing PCR primers?
The melting temperature and sequence specificity to the target.
How are PCR techniques involved in diagnostics of infectious diseases?
They detect and identify pathogens through amplifying their specific DNA targets.
What role does population genetics play in forensic DNA analysis?
It helps establish probabilities of DNA matches based on allele frequencies within a population.
What is the advantage of using a gradient thermal cycler?
It allows for testing multiple annealing temperatures simultaneously to optimize reactions.
What challenge does PCR face when DNA templates are of poor quality?
Degraded DNA can hinder the binding of primers and successful amplification.
What role does PCR play in the analysis of mitochondrial DNA?
It amplifies mtDNA to study maternal lineage and ancestral relationships.
In what ways does PCR speed up the process of gene cloning?
By allowing rapid amplification of gene fragments needed for cloning into vectors.
What is included in the PCR buffer solution and why is it important?
It maintains optimal pH and ionic strength for DNA polymerase activity.
What has been a major controversy regarding patenting PCR technology?
The ownership of PCR techniques and methods has been disputed among various entities.
How is high throughput screening facilitated by PCR technologies?
PCR enables simultaneous testing of multiple samples in a cost-effective manner.
What ethical concerns arise from the use of DNA in forensic databases?
Privacy and the potential misuse of genetic information from individuals.
What does the term 'pharmaceutical drugs in milk' imply in relation to PCR?
It refers to genetically modified animals producing human proteins in their milk for medical use.
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