BEHAVIORAL OBJECTIVES:
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1. To understand DNA replication in the process of mitosis
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2. To develop an understanding of meiosis for the production of germ cells
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3. To understand how proteins are synthesized from codes on the DNA material
The duplication of cells, cell metabolism and cell specialization are also dependent upon the DNA and RNA within the cells’ nuclei.
Mitosis
Cell division known as asexual reproduction takes place in stages and involves a duplication of DNA (double helical) structure in the nucleus before cytoplasmic division occurs.
Annot: The instructor should take the opportunity to use visual aids to demonstrate the differences between mitosis and meiosis and to reinforce that mitosis or cloning is almost a continuous process in most cells. A three dimensional model of the double helical DNA or wall graphs may be used to stress that the DNA/RNA molecules are of prime importance in this study, providing the structure for chromosomes.
Simple asexual division includes the doubling of chromosomes so that each chromosome consists of two chromatids attached in the middle by a centromere. The next phase which is prophase shows each pair of chromatids tightly coiled, visible and paired with its homologue. They form tetrads at the equator of the cell and attach to the spindle. In anaphase the chromatids separate and migrate to opposites ends of the cells. In telophase the cytoplasm pinches off and two new cells are formed called daughter cells. The nuclear membrane disappears after the nucleolus disappears during prophase.
These visible phase changes are the result of some complex molecular biochemistry.
DNA (deoxyribosenucleic acid) consists of a chain of nucleotides, in fact two chains, the chains attached to each other by basis with hydrogen bonding. The double helical structure was determined by several researchers James Watson, Francis Crick and Rosalind Franklin, three of the most eminent.
The importance of the biological function of the DNA in its ability to replicate itself was further investigated by Matthew Meselson and Frank Stahl. Their experiment relied on the ability to differentiate the densities of two isotopes of nitrogen. Using the E. coli bacterial fed on nutrient broth as a source of nitrogen, Crick found that generations replicated DNA with either heavy or light nitrogen determined by the broth.
A nucleotide consists of a 5-carbon sugar called a ribose or deoxyribose, a phosphate and a base. The phosphates link the sugars together to form the outside of a ladder structure and the bases attract complementary bases also attached to sugars and phosphates to form the rungs of the ladder. The four bases are adenine, guanine thymine and cytosine and adenine and thymine always pair as do cytosine and guanine. During chromosome replication, the paired bases separate into two separate strands each one attracting to a new and identical complementary strand.
A number of these bases along the DNA constitute a gene allele therefore replication makes it possible for each new daughter cell in mitosis to have the full genome of a parent cell. In meiosis the germ cells will have only the genes determined by the sequence of bases on one DNA strand.
The sequence of bases in DNA is extremely important in protein synthesis.
The Importance of Bacterium E. coli
Genetic analysis is based on the identification of mutations which are rare DNA changes that disrupt the activity of genes. Since the appearance of mutations is largely a matter of chance, they are most likely to be observed in large populations of organisms which multiply quickly. E. coli is the ideal genetic organism.
E. coli is normal bacterial fauna that inhabit the human colon, where it absorbs digested nutrients. It propagates asexually by binary fission: the chromosome replicates and splits to form two daughter cells. In a sexual phase, mating involves the flow of genetic material through a cytoplasmic bridge called a pillus.
E. Coli is easily grown in a nutrient medium—such as Luria Broth—that contains carbohydrates, amino acids, nucleotide phosphates, salts and vitamins derived from yeast extract and milk protein. Incubation of the E. coli occurs at 37°C (human body temperature).
E. coli bacteria growth falls into the four distinct phases diagramed below. During the logarithmic phase, the culture grows exponentially and cell number doubles every 20-30 minutes. Death rate exceeds fission rate when nutrients are depleted and wastes accumulate.
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Masses of E. coli bacterial cells are grown in a suspension culture and shaking provides the aeration the cells need and keeps cells suspended. Individual colonies can be isolated by spreading the cells on a surface of Luria Broth stiffened with agar. Colonies may be taken off a plate or a stab culture and streaded onto an agar plate using an inoculating loop.
One milliliter of E. coli culture at stationary phase contains approximately one billion cells. A mutation that occurs at a rate of 1/million population should be represented by 1,000 cells. Individual cells are not visible to the naked eye, but colonies of daughter cells are after incubation for 8-12 hours.
E. coli is a prokaryote. It has a single circular chromosome which is unprotected by a nuclear membrane. It contains 5 million base pairs and is therefore only 1/600th the size of the human genome.
Note: Ask students then to compute how many base pairs in the human genome. The chromosomal locations and sequence of a large number of its genes are known.
Because the genetic code is universal, E. coli can accept foreign DNA derived from any organism. A human sequence of DNA inside E. coli is translated and processed in exactly the same manner as the native bacterial DNA.
Hershey experimented with viral phases and determined that it was the DNA which was replicated in bacteria and not the entire phage. Hershey’s experiment also enforced the theory that viral phages have a simplistic structure consisting of a viral protein coat and DNA and that the DNA is the material which enters the bacteria.
Note: Ask students to apply this concept to the diagram of Hershey blender experiment.
(figure available in print form)
(figure available in print form)
(figure available in print form)
(figure available in print form)
(figure available in print form)