The Search for Genetic Material: From Transforming Principle to DNA Proof
The Transforming Principle: Griffith's Experiment (1928)
Researcher: Frederick Griffith performed these experiments in 1928 using the bacterium Streptococcus pneumoniae (also known as Diplococcus pneumoniae).
Experimental Organism Strains:
SIII (Smooth) Strain: This strain is virulent. It synthesizes a polysaccharide capsule that protects itself from the host's immune system. Structurally, it consists of a cell membrane, a cell wall, and an outer polysaccharide layer.
RII (Rough) Strain: This strain is non-virulent. It does not possess a protective capsule and is consequently defeated by the host's immune system.
Experimental Trials and Outcomes:
Trial 1: Live SIII injected into Mice $\rightarrow$ Mice died.
Trial 2: Live RII injected into Mice $\rightarrow$ Mice live.
Trial 3: Heat-killed SIII injected into Mice $\rightarrow$ Mice live.
Trial 4: Heat-killed SIII combined with Live RII injected into Mice $\rightarrow$ Mice died.
Observations from Trial 4: Live S strain bacteria were recovered from the dead mice, indicating that the R strain had transformed into the S strain.
Conclusion: Griffith concluded that this transformation must be due to the transfer of genetic material, which he termed the "Transforming Principle." However, his experiments did not define the biochemical nature of this genetic material.
Biochemical Characterization of the Transforming Principle
Researchers: Oswald Avery, Colin MacLeod, and Maclyn McCarty (1944).
Objective: To determine the chemical nature of the transforming principle and prove that transformation was due to DNA.
Methodology: They used specific enzymes to break down different biochemical components from the heat-killed SIII strain to see which loss prevented transformation.
Experimental Steps:
Protease Treatment: Heat-killed SIII + Live RII + Protease (breaks down protein) $\rightarrow$ Transformation occurred (RII converted to SIII).
RNase Treatment: Heat-killed SIII + Live RII + RNase (breaks down RNA) $\rightarrow$ Transformation occurred (RII converted to SIII).
DNase Treatment: Heat-killed SIII + Live RII + DNase (breaks down DNA) $\rightarrow$ NO Transformation occurred (remained RII).
Key Findings:
Breaking down protein or RNA did not stop transformation.
Only the destruction of DNA prevented the R strain from transforming into the S strain.
Conclusion: The transforming principle is DNA.
The Hershey-Chase Experiment (1952)
Researchers: Alfred Hershey and Martha Chase provided the final, unequivocal proof that DNA is the genetic material.
Experimental Model: They worked with the T2 bacteriophage (a virus that infects bacteria).
Radioactive Labeling: They used isotopes to distinguish between protein and DNA:
Radioactive Sulfur (): Used to label the Protein component (as sulfur is a component of protein but not DNA).
Radioactive Phosphorus (): Used to label the DNA component (as phosphorus is a component of DNA but not protein).
The Process:
Infection: Bacteriophages with either radioactive DNA or radioactive protein were allowed to infect E. coli bacteria.
Blending: The mixture was agitated in a blender to separate the viral protein coats from the bacterial cells.
Centrifugation: The mixture was spun to separate components by weight. The heavier bacteria formed a Pellet, while the lighter viral parts remained in the Supernatant.
Experimental Results (Group 1 - Protein Label):
Radioactivity was detected in the Supernatant (culture medium).
No radioactivity was detected in the Pellet (E. coli cells).
This showed that the protein coat did not enter the bacteria.
Experimental Results (Group 2 - DNA Label):
Radioactivity was detected in the Pellet (E. coli cells).
No radioactivity was detected in the Supernatant.
This showed that viral DNA had entered the bacteria.
Conclusion: DNA is the basic genetic material because it is the substance transferred from the virus to the bacteria to direct the production of new viruses.
Definition of Transduction: Whenever any virus transfers genetic material to bacteria, the phenomenon is called transduction.
Properties of Genetic Material
1. Replication Capability:
The material must be able to replicate itself.
DNA: Yes.
RNA: Yes.
Protein: No.
2. Chemical and Structural Stability:
DNA: More stable and less reactive. It lacks oxygen at a specific position (deoxyribose) and contains the base Thymine (T). This makes it best for the long-term storage of genetic information.
RNA: Less stable and more reactive. Oxygen is present (ribose), and it contains the base Uracil (U). RNA is better suited for the transmission of genetic information.
Base comparison: DNA uses A, G, C, T; RNA uses A, G, C, U.
3. Mutation:
The genetic material must be capable of undergoing slow mutations to allow for evolution.
DNA: Slow evolution.
RNA: Fast evolution.
4. Expression as Mendelian Characters:
The material must be able to express itself through the synthesis of proteins.
Pathway: DNA $\rightarrow$ RNA $\rightarrow$ Protein $\rightarrow$ Enzyme $\rightarrow$ Substrate/Product $\rightarrow$ Mendelian characters (e.g., Height).
Questions & Discussion
Q: Who supported the Griffith effect by providing a molecular explanation?
A: (C) Avery, Maclyn McCarty, and Colin MacLeod.
Q: Identify the correct answer regarding discoveries: 1. Griffith – transformation; 2. Hershey & Chase – transduction.
A: Both 1 and 2 are correct.
Q: Who proved that DNA is the basic genetic material?
A: Hershey and Chase.
Q: The transforming principle of Pneumococcus as found out by Avery, MacLeod, and McCarty was?
A: DNA.
Q: What happened when heat-killed S cells along with R cells were injected into mice?
A: Mice died and showed live S cells in their systems.