BIOL 2161 Practical 3- Experiment involving the use of

radioisotopes

Experiment One
1.

In liquid scintillation counting, cosmic rays, beta particles from

decaying potassium in the glass vial, spontaneous discharges from

the very sensitive photodetectors, and chemicals dissolved in the

scintillation fluid all can contribute to spontaneous flashes of light

that are recorded as counts. The CPM attributable to such sources

are called background. Background counts are often so low relative

to the activity being measured that they are ignored. However if the

number of "real" counts is low, background counts can contribute to

experimental error. It is a good practice to include a vial containing

everything except added radioactivity as a control to determine the

background level. Background CPM are then subtracted directly

from the CPM for the experimental samples.

2.

As a result that each isotopes has its own half-life and emits a

certain pattern of radiation leading to different counting efficiencies.

In this experiment, counting efficiency depends on the energy of the

beta-particle emitted. For instance, a higher counting efficiency

suggests further more beta-particles have passed the solution and

more scintillation mole would be excited which then causes greater

number of photons to be generated. This phenomenon will lead to

an increase of the likelihood that light will be detected resulting a

higher counting efficiency.

Experiment Two
1.

2.

3.

Streptomycin (Str) antibiotic has two effects when existing in the

E.coli cells as a high or low concentration. In a low concentration of

Str medium, it only inhibits growth of E. coli cells by inducing

prokaryotic ribosomes to misread the genetic code. On the other

hand, in a high concentration of Str scenario, it terminates the

initiation of protein synthesis in E.coli cells by binding to 30S subunit

of the E.coli ribosome which could potentially lead to death of E.coli

cells. Additionally, a delay in the inhibition of protein synthesis by

Str perceived from the graph is because synthesis of already

initiated protein is completed.

4.

Since the concentration of E.coli is more diluted by two-fold

compared with the supposed experiment, 6ml of E.coli, the rate of

protein synthesis will also correspondingly decrease by two-fold due

to less E.coli is present to undergo protein synthesis within the same

amount of time taken for 6ml of E.coli in the absence of antibiotics.

5.

According to the first graph in experiment two, penicillin clearly

failed to inhibit E.coli protein synthesis as the trend is still travelling

upwards without a decreasing sign. However, the results do not

indicate penicillin is ineffective antibiotic against protein synthesis

as penicillin aims to penetrate bacterial cell wall which is irrelevant

to protein synthesis.

6.

The comparison between three antibiotics, puromycin, erythromycin

and chloramphenicol against E.coli, can be made based on the

second graph in experiment two. Chloramphenical which shows the

lowest c.p.m count is demonstrated to be one of the two most

effective antibiotics which inhibit protein synthesis in E.coli cells.

Nonetheless, their action against E.coli cells differ from one another.

Chloramphenicol blocks the peptidyl transfer step of elongation on

one of the ribosomal subunit and prevents the aminoacylated end of

charged tRNAs from binding correctly to the A site on the ribosome.

Furthermore, puromycin has a structure similar to that of the 3' end

of a tyrosyl-tRNA which allows it to participate as a substrate in

peptide bond formation, producing peptidyl-puromycin. However,

once puromycin is added to the 3' end of nascent protein, it doesn't

provide a suitable centre for any nucleophilic reactions, and so

protein synthesis is aborted. Finally, the action of erythromycin

blocks the entrance to the exit tunnel and interferes with protein

synthesis. In conclusion, from the graph, it is illustrated that E.coli is

resistant to puromycin and erythromycin but strongly inhibited by

chloramphenicol.