Sect 5 Quiz Easy

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Questions and Answers

What is the role of primers in PCR?

They enhance the stability of the polymerase enzyme.

They are complementary to the 5'-end of the target DNA.

They anneal to the target DNA during the heating process. (correct)

They serve as templates for DNA replication.

At what temperature does the optimal extension of primers occur during PCR?

72 °C (correct)

95 °C

50-60 °C

37 °C

What is the potential downside of using PCR?

It is prone to false-positives due to contamination. (correct)

It can only amplify DNA from genomic libraries.

It requires a large amount of starting DNA.

It cannot be used for cloning purposes.

How does PCR amplify DNA?

Through exponential doublings dependent on the cycles. Signup and view all the answers

What is the purpose of radioactive probes in PCR?

To detect DNA fragments efficiently during hybridization. Signup and view all the answers

What technique is used to detect a specific mRNA within a mixture?

Northern blotting Signup and view all the answers

Which of the following correctly describes Taq polymerase?

It is obtained from Thermus aquaticus and withstands high temperatures. Signup and view all the answers

What is the primary use of cloned DNA fragments in molecular biology?

To study the expression and function of genes. Signup and view all the answers

What is one outcome of nucleotide mutations in DNA?

They can cause alterations at the protein level. Signup and view all the answers

What technique is NOT mentioned as a molecular biological technique?

Gel electrophoresis Signup and view all the answers

How is recombinant DNA technology primarily used?

To identify and clone genes. Signup and view all the answers

Which of the following statements aligns with the Seventh-day Adventist Church's view on human cloning?

Research with animals for cloning purposes is acceptable. Signup and view all the answers

What is one goal of ethical research in the context of human cloning?

To enhance knowledge of life principles. Signup and view all the answers

What potential application is opened by therapeutic cloning?

Development of treatments for diseases. Signup and view all the answers

What is one moral concern regarding human cloning?

It might lead to ethical dilemmas about life creation. Signup and view all the answers

What additional sequences do specific vectors contain besides selectable markers and ori?

CEN and TEL Signup and view all the answers

Which method is indicated for studying RNA molecules?

Northern blot Signup and view all the answers

What is one function of the TEL sequence in vectors?

To protect the ends of linear DNA Signup and view all the answers

What characteristic do shuttle vectors possess?

They can propagate in two different host types. Signup and view all the answers

Which selectable marker is mentioned for yeast in the context of shuttle vectors?

URA3 Signup and view all the answers

What is the primary purpose of creating a genomic library?

To contain every DNA sequence in an organism's genome Signup and view all the answers

How many lambda phages are approximately needed to prepare a haploid human genome library?

150,000 Signup and view all the answers

Which of the following is NOT a component of shuttle vectors?

Telomeric sequences Signup and view all the answers

How long are poly linkers or MCS typically?

50-150 nucleotides Signup and view all the answers

What feature aids in the stable replication of shuttle vectors in yeast?

Yeast centromere (CEN) Signup and view all the answers

What is the direction of DNA synthesis?

5' to 3' Signup and view all the answers

What is the purpose of using a degenerate probe?

To account for redundancy in the genetic code Signup and view all the answers

Which method can NOT cause denaturation of nucleic acids?

Adding water Signup and view all the answers

What is used for detection of an oligonucleotide probe?

Radioactive or fluorescent labels Signup and view all the answers

What breaks hydrogen bonds during nucleic acid denaturation?

Temperature increase Signup and view all the answers

Which base pairing is correct for DNA?

A=T and G≠C Signup and view all the answers

What type of DNA is synthesized when using DNA synthesizers?

Single-stranded DNA Signup and view all the answers

What is the primary purpose of expression vectors derived from plasmids?

To produce abundant quantities of a protein of interest Signup and view all the answers

What is a unique feature of expression vectors mentioned in the content?

They have a promoter fused to the cDNA Signup and view all the answers

In which type of cells can eukaryotic expression vectors be used to express cloned genes?

In yeast and mammalian cells Signup and view all the answers

What is the process of transfection primarily used for?

To introduce cloned genes into cultured animal cells Signup and view all the answers

Which method involves using liposomes to deliver DNA into cells?

Liposome-mediated transfection Signup and view all the answers

What occurs during transient transfection?

The cells lose the plasmid over time Signup and view all the answers

What happens to plasmid vectors used in transient transfection?

They are not incorporated and can be lost Signup and view all the answers

What is a characteristic of eukaryotic expression vectors in relation to prokaryotic systems?

They can function as shuttle vectors for E.coli and mammalian cells Signup and view all the answers

What is produced by disrupting a gene in yeast with a selectable marker?

A heterozygous mutant Signup and view all the answers

What are the consequences of disrupting an essential gene in yeast during sporulation?

Two nonviable haploid spores are produced Signup and view all the answers

What is the purpose of the loxP-Cre recombination system in gene targeting?

To specifically knock out genes in a chosen cell type Signup and view all the answers

How are knockout mice produced in genetic research?

By homologous recombination in embryonic stem cells Signup and view all the answers

What role do embryonic stem (ES) cells play in producing knockout mice?

They provide a source for gene-targeted disruptions Signup and view all the answers

What is the main purpose of creating transgenic animals?

To integrate foreign DNA into their genome Signup and view all the answers

What distinguishes homologous recombination from nonhomologous recombination?

Nonhomologous recombination inserts DNA at random locations Signup and view all the answers

What happens to ES cells with a gene-targeted knockout mutation during selection?

Positive and negative selection methods are used to identify them Signup and view all the answers

Study Notes

Molecular Genetic Techniques

Techniques are used to identify, study, clone, express genes in DNA
Recombinant DNA technology is used to study genes
Mutations in DNA can affect protein function
Techniques like Northern, Southern, Western blots, microarrays, PCR and DNA sequencing are used for research, diagnostics, and forensics
Cloning is the transfer of genetic material by unnatural/artificial means
The Seventh-day Adventist Church views human cloning as unacceptable, but deems animal research acceptable.

Genetic Analysis of Mutations

Strategies for relating genes to function, location and gene-product structure
Classical genetics involves isolating a mutant organism, cloning a gene from a genomic library, or specific amplification of genomic DNA. Once cloned, the encoded protein is produced for analysis. Purified protein can be used to identify a gene
Mutations are alterations in DNA sequences
The different types of mutations include transitions, transversions and indels
Mutations can result in synonymous/silent, missense (conservative/nonconservative) or nonsense substitutions, as well as frameshifts

Haploids and Diploids

A haploid organism has one copy of each chromosome
A diploid organism has two copies of each chromosome and thus two alleles for each gene.
Matching alleles are homozygous, contrasting alleles are heterozygous

Mitosis and Meiosis

Mitosis is cell division in somatic cells
Meiosis is for gamete formation (sex cells such as sperm/egg).

Genetic Screens

Procedures to isolate mutants. Haploid or diploid organism with dominant/recessive screen types.

Conditional Mutations

Mutations with wild-type phenotype under permissive conditions, mutant phenotype in non-permissive conditions (e.g., temperature-sensitive).

Complementation Analysis

Used to determine if mutations on different genes lead to the same phenotype. If mutations are on the same gene, there is no complementation. If on different genes, they complement and have a wild-type phenotype.

Synthetic Lethal Mutations

Phenotypes that exhibit opposite effects from suppression. The deleterious effect of one mutation is worsened by a second mutation(same/related gene). This can reveal whether proteins interact or are redundant.

DNA Cloning and Characterization

DNA cloning is used to make identical copies of DNA molecules.
Recombinant DNA technology joins DNA from different sources.
Restriction endonucleases cut DNA at specific sites, DNA ligase joins fragments.
Restriction enzymes recognize specific (4-8bp) sequences, cleave DNA within the sequence and they are named after the microorganism they are isolated from and are used to cleave/create DNA fragments

Plasmids

Plasmids are extra-chromosomal circular dsDNA, naturally in bacteria and lower eukaryotes. They carry genes for replication, drug-resistance, and polylinkers(multiple cloning sites).
They are used in cloning as vectors for large fragments of DNA.

Other Cloning Vectors

Bacterial, phage (λ, cosmids), and artificial chromosome vectors (BACs/YACS/HACS) with different sizes ideal for different cloning procedures.

DNA libraries

Genomic libraries contain all genes in an organism. cDNA libraries contain only expressed genes.
DNA libraries are prepared by cloning DNA into a vector, then using E. coli to replicate vectors.

Screening DNA Libraries

Methods used to find DNA fragments/clones encoding proteins of interest- detection by amplification (PCR) and by functional complementation in cells.

Blotting Techniques

Southern blotting detects DNA fragments
Northern blotting detects mRNA fragments
Blotting uses electrophoresis, transfer to a membrane and hybridization with a DNA/RNA probe.

In Situ Hybridization

Used for gene expression studies directly in cells/tissues to detect mRNA localization
Detects specific mRNA within a complex mixture
Preserves cellular context.

DNA Microarrays

Used to study gene expression/transcriptional programs in cells under different conditions (can be thousands simultaneous). mRNA is labelled and hybridized to probes.

PCR

A method of amplifying specific DNA sequences using primers. An efficient technique using heat-stable DNA Polymerase.

Reverse Transcriptase PCR (RT-PCR)

Converts mRNA to cDNA, which can then be amplified by PCR. This can be used to measure RNA levels (gene expression)

Expression Vectors

Used to produce a protein from a cDNA clone. Includes a promoter for high transcription and a selectable marker for bacterial selection

Transfection

Process of introducing cloned genes into cultured animal/plant cells
Methods include liposome-mediated transfection and electroporation
Transient transfection means genes are not integrated into chromosome
Stable transfection means genes are incorporated into chromosome

Retroviral Expression

Retroviruses can integrate cDNA into the host cell genome
This allows efficient expression in many cell types as well as production of recombinant retroviral particles

Fusion Proteins

Identifying and Locating Human Disease Genes

Methods to identify disease-causing genes, including mapping. Techniques include, linkage mapping, identifying genetic markers, and analyzing DNA sequences.

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Description

Explore the various molecular genetic techniques used to identify, study, and clone genes. This quiz covers topics such as recombinant DNA technology, mutation analysis, and classic genetics methods for understanding gene functionality. Test your knowledge on techniques including PCR, DNA sequencing, and the implications of cloning.