Chapter 5: Bacterial Transformation & Polymerase Chain Reaction/ Ava Resendez

What is the clinical significance of bacterial transformation in biotechnology?

the core technique for genetic engineering, allowing bacteria to mass-produce crucial human medicines like recombinant insulin, human growth hormone, and erythropoietin, treating diabetes, growth disorders, and anemia

When did Griffith discover transformation?

1928

How did the mouse die In station 1?

The S injected

What did the s-strain contain?

a protective polysaccharide capsule, a sugar-based coat that made it virulent (lethal to mice) by hiding it from the immune system, and also contained the DNA

How was the R-strain different for the S-strain?

the S-strain possessed a protective polysaccharide capsule, making it virulent (disease-causing), while the R-strain lacked this capsule, rendering it harmless (non-virulent), leading to rough-looking colonies in culture compared to the S-strain's smooth colonies

What happens in the station-3 of Griffith experiment?

The heat killed the S-strain and the mouse survives while the bacteria dies.

What happens in station 4?

S&R strain injected in a heat killed strain and the mouse dies.

How did the bacteria demonstrate genetic variation?

through mutations (random DNA changes) and horizontal gene transfer

What happens during DNA release and update in the transformation?

A new bacteria is formed

What happens during DNA integration?

a DNA segment, often from a virus (like retroviruses or HPV) or a transposon, gets inserted into a host cell's genome, facilitated by enzymes (like integrase) that cut and join DNA, leading to permanent genetic change, which can disrupt host genes or activate oncogenes

What happens during bacterial division in the transformation process?

the cell containing the new DNA (often a plasmid) replicates its own chromosome and the foreign DNA, then splits via binary fission, creating two identical daughter cells, each carrying the new genetic information, leading to a colony of transformed cells with the desired trait

What is the clinical significance of bacterial conjugation?

it is the primary mechanism for the rapid spread of antibiotic resistance genes and virulence factors among bacterial populations

What are the different bacterial mating types?

Bacteria don't have "mating types" like sexual organisms

What is the fertility factor?

specific bacterial plasmid (F-factor) enabling gene transfer

What is the conjugation bridge?

a temporary protein channel that forms between two bacteria (a donor and a recipient) during bacterial conjugation, allowing for the direct transfer of genetic material, usually a plasmid

How are genes transferred during conjugation?

through direct cell-to-cell contact

What is high frequency recombination?

describes a special type of bacterium, often E. coli, where the fertility plasmid (F-factor) has integrated into its main chromosome, making it highly efficient at transferring large chunks of its own chromosomal DNA, along with the F-factor, to other bacteria during bacterial conjugation, leading to frequent genetic recombination in the recipient cell

What is bacterial transduction?

the process of transferring bacterial DNA from one bacterium to another via a virus (bacteriophage), acting as a genetic shuttle without direct cell-to-cell contact

What happens during bacteriophage attachment?

the virus (virion) uses its tail fibers to specifically recognize and bind to receptors (like proteins or carbohydrates) on the surface of a susceptible host bacterium, initiating the infection process, which can be reversible or irreversible and leads to either a lytic (killing)

What happens during phage DNA replication?

the bacteriophage injects its genetic material into a host bacterium, hijacks the cell's machinery to copy its DNA and make viral proteins, then assembles new phages, which are released when the host cell lyses (bursts) in the lytic cycle, or the DNA integrates into the host's genome

What happens during packaging and phage release?

During phage packaging, a powerful molecular motor (terminase) uses ATP to pump the viral DNA into a pre-formed, empty protein shell (prohead) through a portal, tightly condensing it

What are the steps of generalized transduction?

(1) Phage infection of donor cell, (2) Host DNA breakdown, (3) Mistaken packaging of bacterial DNA into a phage capsid, (4) Cell lysis releasing transducing particles, (5) Infection of new host, and (6) DNA recombination into the recipient's genome, creating a stable transductant

What happens during prophase integration and excision?

During prophase, especially in meiosis, integration refers to homologous chromosomes pairing up

What is hybrid DNA formation?

the process where two complementary single-stranded nucleic acid (DNA or RNA) molecules bond together to form a double-stranded hybrid structure through specific base pairing

How is DNA transferred to recipient?

through conjugation (direct cell-to-cell contact via pili), transformation (uptake of free DNA from the environment), and transduction (via viruses/bacteriophages)

What is the pre-integration state?

a crucial stage in the lifecycle of retroviruses (like HIV) where viral DNA, newly formed from viral RNA, is packaged with viral and host proteins and transported to the host cell's nucleus for insertion into the host genome

What is the post-integration state?

the final, unified, and fully operational condition of two or more companies after they've merged or been acquired, where their systems, cultures, people, and processes work as one cohesive entity to achieve shared goals and synergies

What is the clinical significance of Bacterial Transduction?

a primary mechanism for the spread of antibiotic resistance genes and virulence factors among bacterial populations

How did genetic engineering revolutionize medicine?

enabling the precise creation of therapeutic proteins (like insulin), developing targeted drugs (like cancer immunotherapies), transforming vaccine production, and offering gene therapies to correct genetic diseases at their source, shifting medicine toward personalized treatments with tools like CRISPR

What is the donor plasmid?

a DNA template, often used in gene editing (like CRISPR), that carries the desired genetic material (like a new gene or correction) to be inserted into a cell's genome

What are restriction enzyme?

proteins from bacteria that cut DNA at very specific short sequences

What are DNA ligase?

a crucial enzyme that acts as "molecular glue," joining DNA strands by forming phosphodiester bonds to seal breaks in the DNA backbone

What do foreign DNA come from?

other organisms, like bacteria, viruses, or other animals

What is a chimera?

a creature or organism composed of cells from two or more distinct individuals

What does CaCl2 solution do to cells?

makes cells, especially bacteria, permeable to foreign DNA by neutralizing negative charges on both the DNA and cell membrane, allowing them to bind

How does the host bacteria reproduce quickly?

binary fission

How can recombinant proteins be used?

as powerful tools and therapeutics in medicine, research, and industry

How did PCR revolutionize medicine?

enabling rapid, sensitive, and precise amplification of DNA

What are primers?

a preparatory coating or base layer used in various fields, like painting (a base coat for paint to stick better), makeup (smooths skin and makes makeup last longer), and molecular biology (a short DNA/RNA strand to start synthesis)

What are dNTP’s?

the fundamental building blocks of DNA, consisting of a deoxyribose sugar, a phosphate group, and one of four nitrogenous bases (Adenine, Cytosine, Guanine, Thymine)

What is Taq DNA polymerase?

a heat-stable enzyme from the bacterium Thermus aquaticus, crucial for the Polymerase Chain Reaction (PCR) to amplify DNA

What are the three steps of PCR?

Denaturation, Annealing, and Extension

What temp does denaturation occur at?

40-60 C

What happens during the denaturation phase?

proteins or nucleic acids (like DNA) lose their specific 3D structure due to stress (heat, pH changes, chemicals), causing them to unfold, break apart hydrogen bonds, and lose function

What happens does annealing phase?

reduces hardness, relieves internal stresses, and increases ductility (softness and flexibility)

What temp does elongation happen at?

72 C

What occurs during elongation stage?

enzymes add building blocks (nucleotides or amino acids) to a growing strand, making it longer

How does PCR multiply?

through repeated cycles of heating and cooling, using denaturation (separating strands), annealing (primers binding), and extension (DNA polymerase synthesizing new strands)

How many cycles of PCR are typical?

20-35