Bacterial Genetics Lab -Crider

Which of the following are potential applications of PCR? 3

-Detection of DNA from victims of massive disasters. -Detection and identification of infectious agents in patient samples. -Detection of DNA during crime scene analysis.

Select each of the true statements about DNA structure 3

-Guanine forms a base pair with cytosine. -Heat can be used to break the hydrogen bonds between nitrogenous bases and separate strands of DNA. -Hydrogen bonds join the nitrogenous bases of nucleotides on opposite strands.

What does the master mix solution contain? 2

-Heat stable polymerase -Free deoxynucleotides

What are the main purposes for adding a loading dye to the samples before they are loaded into the gel? 2

-It contains a colored indicator, allowing migration of the samples to be monitored during electophoresis. -It contains glycerol, which allows the samples to sink into the wells.

What is the importance of creating a master mix? 3

-It ensures more consistent concentrations of reagents for each tube. -It is easier to set up a single tube to add needed components in bulk versus pipetting each sample tube individually. -It helps correct for pipette errors at small volumes.

Which of the following are potential uses of DNA profiles? 4

-Paternity testing -Crime scene investigations -Identifying disaster victims -Tracking a pathogenic microbe during an epidemiologic investigation.

Select the three components of a DNA nucleotide. 3

-Phosphate -Deoxyribose -Nitrogenous base

Which statements correctly explain why DNA from two individuals will produce different fragments after cutting by restriction enzymes? 2

-Single-base differences in their DNA sequences can cause changes in the location of restriction enzyme cutting sites. -The location of restriction enzyme cutting sites determines the size of the DNA fragments when cut with that enzyme.

Which of the following explain the difference in gel results for the outbreak in patient D? 2

-The bacterial DNA sequence is different from the DNA sequence of the outbreak strain. -The outbreak strain had more restriction digest sites than the bacterial DNA in patient D.

Indicate the items needed for PCR: 4

-Thermostable DNA polymerase -Primers -Template DNA with a target sequence -Free deoxynucleotides

What steps were taken to increase the efficiency of the bacterial transformation process? 2

-Treatment with calcium chloride -Brief heat shock

Indicate the four nitrogenous bases that are found in DNA: 4

1. Adenine 2. Thymine 3. Cytosine 4. Guanine

Place the steps of one PCR cycle into the correct order: 3

1. Denaturation 2. Priming 3. Extension

Which two processes are involved in gene expression? 2

1. Transcription 2. Translation

As an epidemiologist, you are collecting statistics on the number of patients that are part of the current Salmonella outbreak. What percentage of these tested patients are positive for the outbreak strain?

20%

Using the DNA ladder provided, estimate the size of the largest DNA fragment in lane 4.

2322 base pairs

What ingredient is often added to media to select for successfully transformed cells?

Antibiotics

What is the first step needed to create a recombinant bacterial cell with a gene of interest?

Creation of a cloning vector

What do restriction enzymes do?

Cut at specific sites that are randomly located throughout a DNA sample.

On what basis does DNA fingerprinting or DNA profiling allow for identifying one cell from another?

Cut sites in DNA are unique to different cells and give different sized fragments

Which of the following best portrays the direction of genetic information flow, according to the central dogma of biology?

DNA ----> RNA -----> Protein

DNA profiles are a quick way of identifying differences in nucleotide sequences. What would be another way to do this even though it would take much longer?

DNA sequencing

Match each step of the PCR cycle with the appropriate description: Double-stranded DNA is separated into single strands by high temperature (95oC).

Denaturation

Match each step of the PCR cycle with the appropriate description: Thermostable DNA polymerase attaches to primers and begins to synthesize new DNA (72oC).

Extension

All RNA molecules serve as a template for protein syntheses. (T/F)

False

Bacterial transformation only occurs in the laboratory when creating recombinant bacteria. (T/F)

False

Because bacteria reproduce asexually they rely on vertical gene transfer to gain genetic diversity. (T/F)

False

Recipient cells successfully transformed with a cloning vector will not typically grow on the selective medium. (T/F)

False

Specific primers are not necessary for PCR to work but they do make the process faster. (T/F)

False

Match each of the items needed for PCR with their function in the reaction: Building blocks that are added to form the new DNA strands.

Free deoxynucleotides

What method is used to separate out restriction fragments after cutting with restriction enzymes?

Gel electrophoresis

What method is used to separate out restriction fragments after cutting with restrictions enzymes?

Gel electrophoresis

Which of the following can be used to detect the products of PCR?

Gel electrophoresis

What is the purpose of the LB agar plate?

It is a non selective medium for bacterial growth

What is the purpose of the LB/amp agar plate?

It is a selective medium containing an antibiotic to inhibit the growth of non-resistant cells.

What is the purpose of the LB/amp/ara agar plate?

It is a selective medium with an antibiotic which prevents growth of non-resistant cells and a sugar that allows for the expression of plasmid gene.

What mutation results in a change in the amino acid sequence?

Missense

Following PCR, the products were loaded into a gel, and electrophoresis was performed. Following staining of the gel, no band of DNA was observed. Based on the observation, was the desired sequence successfully amplified?

NO

Match each of the items needed for PCR with their function in the reaction: Direct the DNA polymerase to the target sequence and provide the starting material for DNA polymerase to add on to.

Primers

Match each step of the PCR cycle with the appropriate description: Cooling to 50-65oC allows primers to attach to template strands of DNA.

Priming (Annealing)

What is the final end product of the Central Dogma of Biology?

Protein

Which of the following is used to cut DNA at specific sequences, prior to the separation of fragments during electrophoresis?

Restriction enzymes

Match each of the items needed for PCR with their function in the reaction: Provides the target sequence which will be amplified.

Template DNA strand

What would be the result of running the gel too long?

The DNA sample will move through the entire gel and exit at the bottom.

Following PCR, the products were loaded into a gel and electrophoresis was performed. Following visualization of the gel, no band of DNA was observed. What explains this result?

The desired sequence was not successfully amplified.

What does EcoRi/Pstl enzyme mixture do?

The enzyme mixture cuts DNA at specific locations, creating various-sized fragments.

What would happen if the electrical current was not left on long enough during gel electrophoresis?

The fragments would not have time to separate very much.

In which of the following ways does the gene for sickle cell hemoglobin differ from the gene for normal hemoglobin?

There is a single nucleotide difference.

Match each of the items needed for PCR with their function in the reaction: Synthesizes new DNA strands.

Thermocycler

Match each of the items needed for PCR with their function in the reaction: Controls and quickly modifies the reaction temperature as needed.

Thermostable heat stable polymerase

What is the purpose of changing micropipette tips after each step?

To prevent cross contamination of the samples

Because PCR is carried our at high temperatures, a heat-stable DNA polymerase is needed. (T/F)

True

Biotechnology is a field with wide-ranging impacts including the legal system. (T/F)

True

Different species of microorganisms as well as different strains within a species can usually be distinguished by genetic differences. (T/F)

True

One base difference in a restriction enzyme site will prevent recognition by the restriction enzyme and cutting at the site. (T/F)

True

PCR is particularly useful when there is very little DNA available for analysis.(T/F)

True

The end product of PCR is amplified DNA.

True

Match the inoculated agar plates with their expected growth after incubation: a) -pGLO on LB: b) -pGLO on LB/amp: c) +pGLO on LB/amp: d) +pGLO on LB/amp/ara:

a) dense white bacterial colonies b) No growth c) White bacterial colonies d) Green fluorescent bacterial colonies

Primers provide the specific location where DNA synthesis will _________.

begin

The use of living organisms and their biological processes to make useful products and address practical problems is

biotechnology

A ___________ ____________ is used to carry genes from another organism into a bacterial cell.

cloning vector

The bonds that join the nucleotides in DNA to form the sugar-phosphate backbone are ______________ bonds.

covalent

The unique pattern of DNA fragment distribution when separated by gel electrophoresis is referred to as a DNA ____________.

fingerprint

human genes can be introduced into bacterial cells via transformation, after which the bacterial cells express the human gene, producing a human protein such as insulin or growth hormone. The bacterial cell is described as _____

genetically-modified and a recombinant cell

Larger fragments migrate _____________ through the agarose gel, compared to smaller fragments.

more slowly

A single nucleotide change in the original sequence of DNA is a

mutation

The visible, observable characteristics of an organism are called its __________________.

phenotype

DNA fragments will migrate through the agarose gel toward the _____________ electrode.

positive

When a gene of interest from one organism is introduced into a plasmid, the plasmid is referred to as a _______________________ plasmid.

recombinant

Plasmids used as cloning vectors often include a gene for antibiotic resistance. This makes it easier to isolate bacterial cells that have been successfully transformed with the plasmid by using a medium containing the same antibiotic. Such an antibiotic-containing medium would be described as

selective

Primers selected for PCR must hybridize with ______________.

the DNA flanking either end of the target sequence.

The process of copying a strand of DNA to form a strand of RNA is called __________.

transcription

Genes for human growth hormone and for human insulin have been placed into plasmids and introduced into bacteria by means of

transformation

The process by which a bacterial cell picks up small pieces of naked DNA from the surrounding environment is referred to as

transformation

The process during which a strand of RNA is used as a template to determine the order of amino acids in the synthesis of a protein is

translation