DNA Isolation and Extraction Techniques

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Questions and Answers

What is the purpose of adding sodium acetate in DNA extraction?

To dissolve the cell membrane and nuclear membrane
To break down peptide bonds and digest contaminated proteins
To precipitate DNA out of the solution (correct)
To stabilize proteins and lipids making up the cell membrane

What is the primary function of Proteinase K in DNA extraction?

To stabilize proteins and lipids making up the cell membrane
To precipitate DNA out of the solution
To digest contaminated proteins and break down peptide bonds (correct)
To break down the cell membrane and nuclear membrane

What is the composition of the supernatant after centrifugation in DNA extraction?

Contains nucleic acids and water-soluble proteins (correct)
Contains cell debris and insoluble proteins
Contains nucleic acids and membrane lipids
Contains only DNA molecules

What is the purpose of grinding or sonication in DNA extraction?

To mechanically break down cells and release DNA (D) Signup and view all the answers

What is the function of SDS in DNA extraction?

To dissolve the cell membrane and nuclear membrane (A) Signup and view all the answers

What is the purpose of centrifugation in DNA extraction?

To separate debris from soluble components (A) Signup and view all the answers

In which part of the cell is DNA typically found?

Nucleus (A) Signup and view all the answers

What is the purpose of cell lysis in DNA extraction?

To release DNA from the cell (B) Signup and view all the answers

What is the primary function of plasmids in bacterial cells?

To carry genes associated with specialized functions (D) Signup and view all the answers

What is the term for the size range of bacterial genomes?

50kb - 13mn (C) Signup and view all the answers

What is the purpose of an operon in bacteria?

To allow for simultaneous regulation of gene clusters (A) Signup and view all the answers

What is the result of lactose binding to the repressor in the lac operon?

The repressor changes conformation and allows RNA polymerase to transcribe (D) Signup and view all the answers

What is the function of the permease enzyme in the lac operon?

To catalyze transport of lactose into the cell (B) Signup and view all the answers

What is the term for the promoter, operator, and structural genes in an operon?

Functional unit of DNA (B) Signup and view all the answers

What is the purpose of plasmids in Pseudomonas species?

To carry genes associated with specialized functions, such as degradation of camphor and toluene (A) Signup and view all the answers

What is the result of a small genome size in bacterial species?

A restricted ecological niche (A) Signup and view all the answers

What is the purpose of DNA replication in cell division?

To ensure that each daughter cell receives an exact copy of the parental DNA (C) Signup and view all the answers

What is the primary function of the RNA polymerase enzyme during transcription?

To synthesize a complementary mRNA molecule (B) Signup and view all the answers

What is the significance of Chargaff's ratio in understanding DNA structure?

It provides evidence for the double helix model (C) Signup and view all the answers

What is the role of promoters in the transcription process?

They initiate the transcription process (C) Signup and view all the answers

What is the result of horizontal gene transfer between cells of the same generation?

The exchange of genetic information between cells (A) Signup and view all the answers

What is the contribution of Rosalind Franklin's work to the discovery of DNA structure?

Her work provided the data for the helical structure of DNA (C) Signup and view all the answers

What is the biological problem that DNA replication solves?

The issue of where a cell's DNA goes when the cell divides (C) Signup and view all the answers

What is the outcome of the process of DNA replication?

The production of two identical daughter cells (A) Signup and view all the answers

What is the purpose of adding a volume of sterile nuclease free water to the microfuge tube?

To achieve a final reaction volume of 20 microliters (D) Signup and view all the answers

What is the definition of one unit of restriction enzyme?

1 microgram of DNA digested in 1 hour at 37°C (D) Signup and view all the answers

Why is it important to incubate the reaction mixture at 65°C after the restriction digestion?

To inactivate the restriction enzyme (A) Signup and view all the answers

What is the consequence of prolonged incubation times during the restriction digestion reaction?

Star activity (D) Signup and view all the answers

What is the purpose of running the DNA in an agarose gel after the restriction digestion?

To ensure that the digest was successful (A) Signup and view all the answers

What is the term for the DNA segment that shows the positions of the restriction sites?

Restriction map (D) Signup and view all the answers

What is the advantage of using control DNA with known restriction sites?

To test the enzyme activity (B) Signup and view all the answers

What is the purpose of screening samples using restriction digestion?

To identify samples diagnostically (A) Signup and view all the answers

What is the purpose of adding salt to the DNA sample during DNA extraction?

To stabilize the negatively charged backbone (B) Signup and view all the answers

What is the result of cutting a plasmid with restriction enzymes?

A linear piece of DNA (A) Signup and view all the answers

What is the purpose of adding a nucleotide stain to the agarose mixture during gel preparation?

To visualize the DNA fragments during electrophoresis (D) Signup and view all the answers

Why do smaller DNA fragments move more quickly during electrophoresis?

Because they are shorter in length (B) Signup and view all the answers

What is the purpose of the DNA ladder in DNA gel electrophoresis?

To provide a standard against which to compare the DNA fragments (A) Signup and view all the answers

What is the direction of DNA migration during electrophoresis?

From the cathode to the anode (D) Signup and view all the answers

What is the purpose of using a combination of restriction enzymes in DNA fingerprinting?

To produce distinct fragments for each individual (C) Signup and view all the answers

What is the result of adding ice-cold alcohol to the DNA sample during DNA extraction?

The DNA precipitates out of the solution (A) Signup and view all the answers

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Study Notes

Plasmids

Plasmids are extrachromosomal, autonomous, and self-replicating circular DNA molecules
They contain genes necessary for replication and may integrate into the chromosome
Plasmids carry genes associated with specialized functions, while chromosomes carry genes essential for bacterial growth (housekeeping genes)
Examples of metabolic activities determined by plasmids: degradation of camphor, toluene, serine, and salicylic acid by Pseudomonas s.p.

Genome Size

Genome size ranges from 50kb to 13 million base pairs
Smaller genomes are often found in species with restricted ecological niches (e.g., obligate intracellular parasites, endosymbionts)

Operons

Operons are functioning units of DNA containing a cluster of genes under the control of a single promoter
Allows for simultaneous regulation of gene clusters as genes share a common promoter and repressor
Example: Lac operon contains a promoter, operator, and structural genes (β-galactosidase, β-galactoside permease, and β-galactoside transacetylase)

DNA Structure and Replication

DNA is a linear polymer of 4 types of bases linked by sugar-phosphate groups
The ratio of bases is always A=T, G=C (Chargaff's ratio)
DNA is helical with a known helix width and base spacing
Watson and Crick published a model of DNA in 1953 using X-ray crystallography data from Rosalind Franklin

DNA Isolation and Restriction Enzyme Analysis

DNA is isolated from cells using cell lysis, protein removal, and DNA recovery
Restriction enzymes are used to cut DNA at specific recognition sites
Different types of DNA extraction are used for different cell types
Applications of DNA isolation and restriction enzyme analysis include DNA fingerprinting and recombinant DNA technology

DNA Fingerprinting

DNA fingerprinting can identify novel patterns of DNA specific to an individual
Involves the use of restriction enzymes that interact with DNA and recognize specific DNA sequences
Different restriction enzymes recognize different DNA sequences, resulting in distinct fragments
DNA fragments can be examined using DNA gel electrophoresis

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