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Questions and Answers
What is the purpose of using adaptors in molecular cloning?
What is the purpose of using adaptors in molecular cloning?
- To increase the size of plasmids
- To promote the transformation of bacterial cells
- To join different types of molecule ends (correct)
- To enhance the stability of DNA molecules
What occurs during the brief heat shock in the transformation of E.coli?
What occurs during the brief heat shock in the transformation of E.coli?
- The plasmids replicate within the cell
- The DNA is completely degraded
- The cell membrane becomes permeable to DNA (correct)
- The E.coli undergoes cell division
What is the role of selectable markers in the transformation process?
What is the role of selectable markers in the transformation process?
- To inhibit cell division in untransformed cells
- To determine vector size
- To facilitate gene expression
- To identify successfully transformed cells (correct)
What happens when target DNA is inserted into the Tetr gene of a vector like pBR322?
What happens when target DNA is inserted into the Tetr gene of a vector like pBR322?
Which method is used to introduce vector DNA into host cells?
Which method is used to introduce vector DNA into host cells?
During transformation, which of the following is NOT a characteristic of untransformed host cells?
During transformation, which of the following is NOT a characteristic of untransformed host cells?
In the context of insertional selection, what does the presence of the ampr gene indicate?
In the context of insertional selection, what does the presence of the ampr gene indicate?
Which of the following best describes transfection?
Which of the following best describes transfection?
Which restriction enzyme is NOT present at the promoter of the tetracycline gene?
Which restriction enzyme is NOT present at the promoter of the tetracycline gene?
What is the expected outcome when purified DNA is cut with Bam H1 and inserted into a phage vector?
What is the expected outcome when purified DNA is cut with Bam H1 and inserted into a phage vector?
Which type of vector contains two restriction sites flanking a replacement region?
Which type of vector contains two restriction sites flanking a replacement region?
What is the main advantage of using phage vectors over plasmid vectors for cloning?
What is the main advantage of using phage vectors over plasmid vectors for cloning?
Which component is NOT a part of phage λ?
Which component is NOT a part of phage λ?
In the lytic cycle, how does multiplication of the virus take place?
In the lytic cycle, how does multiplication of the virus take place?
What type of phage vector is represented by Lambda gt 10?
What type of phage vector is represented by Lambda gt 10?
What is a distinctive feature of a prophage?
What is a distinctive feature of a prophage?
What characterizes monoclonal antibodies?
What characterizes monoclonal antibodies?
What is the primary purpose of using monoclonal antibodies in laboratory tests?
What is the primary purpose of using monoclonal antibodies in laboratory tests?
Which of the following steps is NOT part of the hybridoma technology process?
Which of the following steps is NOT part of the hybridoma technology process?
In genetic engineering, what role do cloning vectors play?
In genetic engineering, what role do cloning vectors play?
Which enzyme is associated with hypoxanthine guanine phosphoribosyl transferase (HGPRT) activity?
Which enzyme is associated with hypoxanthine guanine phosphoribosyl transferase (HGPRT) activity?
What is the outcome of fusing B-lymphocytes with myeloma cells?
What is the outcome of fusing B-lymphocytes with myeloma cells?
Which of the following uses of monoclonal antibodies does NOT pertain to diagnostics?
Which of the following uses of monoclonal antibodies does NOT pertain to diagnostics?
What is the term used for the process of making recombinant DNA?
What is the term used for the process of making recombinant DNA?
What is the primary function of the λ part in cosmids?
What is the primary function of the λ part in cosmids?
How are YACs different from typical plasmid vectors?
How are YACs different from typical plasmid vectors?
Which feature of cosmids helps in the selection of cells containing the vector?
Which feature of cosmids helps in the selection of cells containing the vector?
What allows phage vectors to identify bacterial cells effectively?
What allows phage vectors to identify bacterial cells effectively?
Which component is NOT typically part of a YAC?
Which component is NOT typically part of a YAC?
What determines the stability of linear YAC in yeast cells?
What determines the stability of linear YAC in yeast cells?
How does a virus insert genetic material into a host cell?
How does a virus insert genetic material into a host cell?
What is the maximum amount of DNA a cosmid can accommodate?
What is the maximum amount of DNA a cosmid can accommodate?
What is a primary characteristic of continuous cell lines?
What is a primary characteristic of continuous cell lines?
Which of the following processes is primarily used to produce cell suspensions for primary cell cultures?
Which of the following processes is primarily used to produce cell suspensions for primary cell cultures?
What is the role of DNA and RNA probes in microbiology?
What is the role of DNA and RNA probes in microbiology?
What type of cell culture is characterized by multiple generations and indefinite growth?
What type of cell culture is characterized by multiple generations and indefinite growth?
In genetic engineering, what is one application of producing transgenic animals?
In genetic engineering, what is one application of producing transgenic animals?
What is the primary purpose of using artificial vaccines?
What is the primary purpose of using artificial vaccines?
Which method is commonly used in the detoxification of industrial effluents?
Which method is commonly used in the detoxification of industrial effluents?
What is one of the features of finite cell lines?
What is one of the features of finite cell lines?
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Study Notes
DNA and RNA Probes
- Utilized for diagnosing diseases like kala azar, sleeping sickness, and malaria.
Artificial Vaccines
- Developed to stimulate immune response against specific pathogens.
Gene Therapy
- A method for treating genetic diseases by correcting defective genes.
DNA Fingerprinting
- A technique used for personal identification and forensic analysis.
Scope and Importance in Microbiology
- Biochemical production including alcohol and antibiotics.
- Protein and enzyme remodeling to enhance functionality.
- Control of plant diseases and pest management (viruses, fungi, bacteria).
- Industrial effluent and sewage treatment for detoxification.
Genetic Engineering
- Involves manipulating genetic material to produce transgenic animals.
- Enables rapid clonal multiplication of crops for agricultural improvement.
- Focuses on developing virus-resistant and disease-resistant plant varieties.
Fundamentals of Animal Cell Culture and Hybridoma Technology
- Cell culture allows for controlled cell growth in nutrient media.
- Primary cell culture derived from directly isolated tissues.
- Disaggregation techniques include enzymatic digestion and physical disruption.
- Secondary culture involves subculturing from primary cultures.
Cell Lines
- Defined as permanently established cell cultures with indefinite division potential.
- Finite cell lines have limited division capability and show contact inhibition.
- Continuous cell lines (transformed/cancerous) grow indefinitely, lacking density limitations.
Monoclonal Antibodies (mAb)
- Produced from a single clone of B-cell, specific to a singular antigen.
- Used for detecting antibodies, purifying substances, and diagnosing diseases.
- Essential in immunohistochemistry and serological tests.
Hybridoma Technology Steps
- Immunization of a rabbit leads to antibody production.
- Fusion of B-lymphocytes with myeloma cells to produce hybrid cells.
- Selected hybridoma cells are cultured for large-scale mAb production.
Gene Cloning and DNA Sequencing
- Involves isolation and combination of DNA fragments for replication.
- Requires cloning vectors (plasmid or phage) for DNA insertion and host transfer.
Phages
- Bacterial-infecting viruses following lytic or lysogenic cycles.
- Examples include λ phage and M13 with specific integration capabilities.
Vectors in Genetic Engineering
- Phage vectors more efficient for large DNA cloning (20-23 kb).
- Plasmid vectors accommodate less than 10 kb and require special introduction methods.
- Cosmids combine features of plasmids and phage, accommodating large DNA inserts (up to 45 kb).
Yeast Artificial Chromosome (YAC)
- Linear DNA segments that mimic yeast chromosomes, capable of cloning significant DNA (up to 1 Mb).
- Contains necessary elements for replication, consists of left and right arms with telomeres.
Transformation and Transfection Methods
- Transformation involves adding new DNA to bacterial cells, using calcium chloride and heat shock.
- Transfection includes phage particles packaging recombinant DNA for infection into bacteria.
Detection of Recombinant Molecules
- Selection methods utilize antibiotic resistance as a marker to distinguish transformed hosts.
- Insertional selection inactivates certain antibiotic resistance genes to ensure accurate identification of recombinants.
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