9.1

b. Which tube number served as a negative control in these experiments?

1

1. To fulfill its role, the genetic material must meet what four criterial listed in the reading?

1. Information: The genetic material must contain the information necessary to construct an entire organism. In other words, it must provide the blueprint for determining the inherited traits of an organism. 2. Transmission: During reproduction, the genetic material must be passed from parents to offspring. 3. Replication: Because the genetic material is passed from parents to offspring, and from mother cell to daughter cells during cell division, it must be copied. 4. Variation: Within any species, a significant amount of phenotypic variability occurs. For example, Mendel studied several characteristics in pea plants that varied among different strains.

1. Describe the four criteria that the genetic material must fulfill.

1. Information: The genetic material must contain the information necessary to construct an entire organism. In other words, it must provide the blueprint for determining the inherited traits of an organism. 2. Transmission: During reproduction, the genetic material must be passed from parents to offspring. 3. Replication: Because the genetic material is passed from parents to offspring, and from mother cell to daughter cells during cell division, it must be copied. 4. Variation: Within any species, a significant amount of phenotypic variability occurs. For example, Mendel studied several characteristics in pea plants that varied among different strains. These included height (tall versus dwarf) .

2. Analyze the results of (1) Griffith, (2) Avery, MacLeod, and McCarty, and (3) Hershey and Chase, and explain how they indicated that DNA is the genetic material.

1. fig 9.1 Griffith looked at S. pneumonia if smooth=was coated so escaped being killed by mouse immune system-mouse was dead, live R type- mouse alive, S dead -mouse alive, If S dead + R alive- mouse died. Moreover S alive was found in tissue. He concluded that there must be something transformed from S dead to R type, and converted the R type to S alive. Avery, Colin MacLeod, and Maclyn McCarty realized that Griffith's observations could be used as part of an experimental strategy to identify the genetic material. They asked the question, What substance is being transferred from the dead type S bacteria to the live type R? To separate these components and to determine if any of them was the genetic material, Avery, MacLeod, and McCarty used established biochemical purification procedures and prepared bacterial extracts from type S strains containing each type of these molecules. After many repeated attempts with different types of extracts, they discovered that only one of the extracts, namely, the one that contained purified DNA from type S bacteria, was able to convert the type R bacteria into type S. As shown in Figure 9.2, when this extract was mixed with type R bacteria, some of the bacteria were converted to type S. However, if no DNA extract was added, no type S bacterial colonies were observed on the petri plates. To further verify that the DNA in the extract was indeed responsible for the transformation, Avery, MacLeod, and McCarty treated samples of the DNA extract with enzymes that digest DNA (called DNase), RNA (RNase), or protein (protease) (see Figure 9.2). When the DNA extracts were treated with RNase or protease, they still converted type R bacteria into type S. These results indicated that any RNA or protein in the extract was not acting as the genetic material. However, when the extract was treated with DNase, it lost its ability to convert type R into type S bacteria. These results indicated that the degradation of the DNA in the extract by DNase prevented conversion of type R to type S. This interpretation is consistent with the hypothesis that DNA is the genetic material. A more elegant way of saying this is that "DNA is the transforming principle." Last exp fig 7.5 involves bacteriophage T2 .newly made phages were radiolabeled with 35 S or 32 P. In the experiment described in Figure 9.5, Hershey and Chase began with E. coli cells, 35 S-labeled phage, and 32 P-labeled phage. In one flask, E. coli cells were mixed with 35 S-labeled phage and, in the other flask, the cells were mixed with 32 P-labeled phage. The phages were given sufficient time to inject their genetic material into the cells, and then samples were taken from each flask and subjected to a shearing force using a blender. This treatment removed the phage coats from the surface of the bacterial cells. The samples were then subjected to centrifugation at a speed that caused the heavier bacterial cells to form a pellet at the bottom of the tube, whereas the lighter phage coats remained in the supernatant, the liquid found above the pellet. The amount of radioactivity in the supernatant (emitted from either 35 S or 32 P) was determined using a Geiger counter. As seen in The Data, most of the 35 S isotope was found in the supernatant. Because the shearing force was expected to remove the phage coat, this result indicates that the empty phages contain primarily protein. By comparison, only about 35% of the 32 P was found in the supernatant following shearing. Therefore, most of the DNA was located within the bacterial cells in the pellet. These results are consistent with the idea that DNA is injected into the bacterial cytoplasm during infection, which would be the expected result if DNA is the genetic material.

C3. Look up the meaning of the word transformation in a dictionary and explain whether it is an appropriate word to describe the transfer of genetic material from one organism to another.

3. It enters the cytoplasm.

6. What percentage of 32-P remained in the intact E. coli cells? Just enter the number

65% bcs 100-35%

DNA

Abbreviation for Deoxyribonucleic Acid

RNA

Abbreviation for ribonucleic acid

DNase

An enzyme that catalyzes the hydrolysis of DNA polynucleotide strands into nucleotides.

RNase

An enzyme that catalyzes the hydrolysis of RNA polynucleotide strands into nucleotides.

2. In the Hershey and Chase experiment involving T2 phage, after blender treatment and centrifugation a. most of the 32 P was in the supernatant and most of the 35 S was in the pellet. b. most of the 35 S was in the supernatant and most of the 32 P was in the pellet. c. equal amounts of 32 P and 35 S were in the supernatant and pellet. d. none of the above was observed.

B

C1. What is the meaning of the term genetic material?

C1. Answer: The term genetic material refers to the actual substance that contains genetic information. It is usually DNA, but in some viruses it can be RNA.

e. What did the results of these experiments indicate was the transforming factor? (one word)

DNA

c. Which one of the experimental treatments resulted in a lack of transformed cells (i.e. what was added to the tube with type S DNA extract)? (One word)

DNase

E1. Genetic material acts as a blueprint for an organism's traits. Explain how Griffith's experiments indicated that genetic material was being transferred to the type R bacteria.

E1. Answer: A trait of pneumococci is the ability to synthesize a capsule. There needs to be a blueprint for this ability. The blueprint for capsule formation was being transferred from the type S to the type R bacteria. (Note: At the molecular level, the blueprint is a group of genes that encode enzymes that can synthesize a capsule.)

E2. With regard to the experiment described in Figure 9.2, answer the following: A. List several possible reasons why only a small percentage of the type R bacteria was converted to type S. B. Explain why an antibody was used to remove the bacteria that are not transformed. What would the results look like, in all five cases, if the antibody/centrifugation step had not been included in the experimental procedure? C. The DNA extract was treated with DNase, RNase, or protease. Why was this done? (In other words, what were the researchers trying to demonstrate?)

E2. Answer: A. There are different possible reasons why most of the cells were not transformed. 1. Most of the cells did not take up any of the type S DNA. 2. The type S DNA was usually degraded after it entered the type R bacteria. 3. The type S DNA was usually not expressed in the type R bacteria. B. The antibody/centrifugation steps were used to remove the bacteria that had not been transformed. It enabled the researchers to determine the phenotype of the bacteria that had been transformed. If this step was omitted, there would have been so many colonies on the plate it would have been difficult to identify any transformed bacterial colonies, because they would have represented a very small proportion of the total number of bacterial colonies. C. They were trying to demonstrate that it was really the DNA in their DNA extract that was the genetic material. It was possible that the extract was not entirely pure and could contain contaminating RNA or protein. However, treatment with RNase and protease did not prevent transformation, indicating that RNA and protein were not the genetic material. In contrast, treatment with DNase blocked transformation, confirming that DNA is the genetic material.

3. In figure 9.1, explain why the mouse in part (d) died.

FIGURE 9.1 Answer: In this experiment, the type R bacteria had taken up genetic material from the heat-killed type S bacteria, which converted the type R bacteria into type S. This enabled them to proliferate within the mouse and kill it

7. True or False: The injection of live type R bacteria caused mice to die

False

Types R and S

He then observed whether or not the bacteria caused a lethal infection. Griffith was working with two strains of S. pneumoniae, a type S (S for smooth) and a type R (R for rough). When injected into a live mouse, the type S bacteria proliferated within the mouse's bloodstream and ultimately killed the mouse (Figure 9.1a). Following the death of the mouse, Griffith found many type S bacteria within the mouse's blood. In contrast, when type R bacteria were injected into a mouse, the mouse lived (Figure 9.1b). To verify that the proliferation of the smooth bacteria was causing the death of the mouse, Griffith killed the smooth bacteria with heat treatment before injecting them into the mouse. In this case, the mouse also survived. When streaked onto petri plates containing a solid growth medium, capsule-secreting strains have a smooth colony morphology, whereas those strains unable to secrete a capsule have a rough appearance. The different forms of S. pneumoniae also affect their virulence, or ability to cause disease. When smooth strains of S. pneumoniae infect a mouse, the capsule allows the bacteria to escape attack by the mouse's immune system. As a result, the bacteria can grow and eventually kill the mouse. In contrast, the nonencapsulated (rough) bacteria are destroyed by the animal's immune system.

4. In the Hershey and Chase experiments, why were radioactive sulfur and phosphorus used?

Hershey and Chase used radioisotopes to distinguish proteins from DNA. Sulfur atoms are found in proteins but not in DNA, whereas phosphorus atoms are found in DNA but not in phage proteins. Therefore, 35 S (a radioisotope of sulfur) and 32 P (a radioisotope of phosphorus) were used to specifically label phage proteins and DNA, respectively.

3. Use the terms listed below to correctly explain concepts, assigned figures, and specified end-of-chapter questions. Molecular genetics DNA RNA Griffith Streptococcus pneumoniae Types R and S Transformation Avery, MacLeod, McCarty DNase RNase Proteases T2 bacteriophage Hershey and Chase Radioactive sulfur Radioactive phosphorus

Molecular genetics is the study of DNA function and structure at molecular level. DNA is the deoxyribonucleic acid, RNA is ribonucleic acid. Griffith used two types of S. pneumonia called types R and S. S type contained a capsule and when alive S injected in mice it caused death in mice. When alive type R was injected in mice they survived (rough no coat). When dead S and alive R were injected mice died. Griffith realized that there should be a substance that is transmitted from dead S to R type that converts R to S. He called this process transformation. Avery MacLeod, McCarty wanted to find out what was the substance that was transmitted. They plated on petri dish strains of S type. When R type was cultured no colonies were produced. When S type purified was mixed with R type, there was colonies. Furthermore they added three types of enzymes to see if DNA indeed was responsible for genetic information. When Type R and S type extract was treated with protease or with RNase the genetic material was still transmitted proving that they didn't influence genetic material transmission. However when DNase enzyme was introduced, which cut the DNA, no colonies were produced due to lack of DNA. Hershey and Chase also conducted an experiment to prove that DNA was carrying the genetic material. They used T2 bacteriophage. Radioactive sulfur was used to radiolabel the protein, and radioactive phosphorus was used to label DNA. After the two were mixed in blender and centrifuged, the protein coat mostly was sitting at the surface, while the DNA was heavier and was noticed at the bottom. This illustrates that DNA was introduced in cytoplasm while the phage remained outside the bacteria.

d. What two substances were the most likely possible contaminants of the Type S DNA extract (one word each separated by one space)?

RNA protein

Griffith

Scientist that worked with strains of Streptococcus to establish the principle of transformation

Hershey and Chase

Scientists that used differential radioactive labeling of T2 bacteriophage to contribute evidence that DNA is the genetic material.

streptococcus pneumoniae

Strain of bacteria that Griffith worked with on his transformation experiments

Transformation

Term coined by Griffith to explain how R strain bacteria could be changed to S strain bacteria.

2. What is meant by the term "virulence"? In regards to Griffith's work, which strain of S. pneumoniae was virulent to mice?

The different forms of S. pneumoniae also affect their virulence, or ability to cause disease. When smooth strains of S. pneumoniae infect a mouse, the capsule allows the bacteria to escape attack by the mouse's immune system. As a result, the bacteria can grow and eventually kill the mouse. In contrast, the nonencapsulated (rough) bacteria are destroyed by the animal's immune system.

molecular genetics

The study of DNA structure and function at the molecular level.

Avery, MacLeod, McCarty

Three scientists that provided additional evidence that DNA was the transfoming factor. Used type S extract exposed to various agents to see if it would still transform the type R bacterial cell

T2 bacteriophage

Type of bacteriophage used by Hershey and Chase to provide additional evidence that DNA is the transforming factor.

Protease

Type of enzyme that catalyzes the hydrolysis of protein chains into amino acids.

1. Consider the figure representing a summary of the experiments of Avery, MacLeod, and McCarty. a. What was added to the tubes with type R cells and type S extract to aggregate type R cells that were not transformed? (One word)

antibodies

2. What item (one word) was used to shear the phage coats from the surface of the bacterial cells at determined times?

blender

5. What lab technique was used to separate the bacterial cells from the phage coats? One word

centrifugation

1. In the experiment of Avery, McLeod, and McCarty, the addition of RNase and protease to the DNA extracts a. prevented the conversion of type S bacteria into type R bacteria. b. allowed the conversion of type S bacteria into type R bacteria. c. prevented the conversion of type R bacteria into type S bacteria. d. allowed the conversion of type R bacteria into type S bacteria.

d

2. Which of the four criteria for the genetic material involves the need to encode instructions for determining the inherited traits of an organism?

information

4. Which of the isotopes above is found in DNA

phosphorus

1. The head of the bacteriophage is composed of ________ and ______ is contained inside the capsid head. One word each blank and separate them by one space

proteins DNA

Radioactive phosphorus

radioisotope used to distinguish proteins from DNA. Phosphorus atoms are found in DNA but not in phage proteins

Radioactive sulfur

radioisotope used to distinguish proteins from DNA. Sulfur atoms are found in proteins but not in DNA

1. Which of the four criteria for the genetic material involves the need to produce sister chromatids prior to meiosis?

replication

5. The presence or absence of a polysaccharide capsule lead F. Griffith to designate two strains of S. pneumoniae as ______ and ______ (one word each separated by one space).

smooth rough

3. What two radioisotopes were used in labeling the phase? Just write the names of the elements separated by one space.

sulfur phosphorus

6. What term (one word) did Griffith use to describe the process of the dead type S bacteria converting the live type R bacteria into type S bacteria?

transformation

4. Which of the four criteria for the genetic material involves the need for passing genetic information from parents to offspring?

transmission

3. Which of the four criteria for the genetic material involves the need for the possibility of phenotype differences within species?

variation