MICROBIAL GENETICS Primary Structure of RNA

Genetics
- A study of heredity
- How the information contained in Nucleic Acids is expressed?
- How is this type of molecule duplicated?
- How are these duplicated molecules transmitted to progeny?

Nucleic Acids
- Are large organic molecules that are found in all cells.
Two Types:
o DNA [Deoxyribonucleic Acid] – directs protein production.
o RNA [Ribonucleic Acid]
Composition:
o Constructed from a string of small molecules called nucleotides.
Components of a Nucleotide

Nitrogenous Base
Primary Structure of DNA DNA and RNA

Ribonucleic Acids Deoxyribonucleic Acids

- Normally single stranded molecules. - Determines the characteristics of an organism and maintains and controls the
Types [based on their function] vital processes of all cells.
o mRNA [messenger] How is genetic information expressed?
o tRNA [transfer] o Transcription [involves formation of RNA molecules using DNA as a
o rRNA [ribosomal] template]
o Translation [consists of the synthesis of a protein using the genetic
information in the RNA]
Transcription of DNA to RNA DNA Replication in Bacteria
- Genome – total genetic information in bacteria which consists of circular DNA
molecules found within the cell.
o Most of the genome is contained in a single bacterial chromosome,
although smaller pieces of circular DNA called plasmids may also
carry a few important genes such as those coding for resistance to
microbial drugs.

RNA Synthesis in Bacteria

- Transcription
o Involves the assembly of nucleotides by an enzyme called RNA
polymerase that uses a strand of DNA as its template.
o Begins when RNA polymerase binds to the DNA at the promoter site
near the gene to be transcribed.
o RNA polymerase travels along the length of the DNA strand until it
reaches a termination site.
o After mRNA is made, it will be used as a guide to make proteins.
o Ribosomal RNA, after it is made, becomes associated with proteins to
form ribosomes.
o tRNA are small RNA molecules that are involved in translating the
The Central Dogma information in the mRNA into proteins.
- Gene Mutagen
o The unit of genetic information or hereditary material contained in - Mutation relevant
DNA molecule, - Cause DNA damage that can be converted to mutations.
o Sequences nucleotide in the DNA molecule that codes for RNA
molecule and ultimately for they synthesis of a protein. Physical mutagens
- Reverse Transcription High-energy ionizing radiation: X-rays and g-rays -> strand breaks and
o Example: Certain cancer viruses [retroviruses] can synthesize DNA base/sugar destruction
using RNA as a template. Non-ionizing radiation: UV light -> pyrimidine dimers

Chemical mutagens
Base analogs: direct mutagenesis; “mimic” certain bases but pair with others
[e.g. 5-fluoracil, cytarabine]
Nitrous acid: determinates C to produce U
Alkylating agents
Intercalating agents
Base pair altering chemicals: deaminates like nitrous acid, nitrosoguanidine,
or alkylating agents like Cytoxan.
Mutation: Ionizing Radiation of the F plasmid is transferred from the F+ to the F-, both cells are
- Ionizing radiation [X-rays, gamma rays, UV light] causes the formation of ions now F+
that can react with nucleotides and the deoxyribose-phosphate backbone.
- Nucleotide excision repairs mutatuons. 2. Transduction: use of bacteriophage [bacterial virus] to transfer DNA between
cells.
Genetic Transfer o The process of moving bacterial DNA from one cell to another using a
- Horizontal gene transfer among prokaryotes bacteriophage.
o Horizontal gene transfer o Bacteriophage are bacterial viruses that consists of a small piece of
▪ Donor cell contributes part of genome to recipient cell DNA inside a protein coat, which binds to the bacterial surface, then
Three Types: injects the phage DNA; then the phage DNA takes over the cell’s
o Transformation machinery and replicates many virus particles.
o Transduction
o Bacterial conjugation Two forms of transduction:
a. Generalized: any piece of the bacterial genome can be transferred
Bacterial Sexual Processes b. Specialized: only specific [pieces of the chromosome can be
- Eukaryotes have the transferred.
processes of meiosis to
reduce diploids to haploids, 3. Transformation: naked DNA is taken up from the environment by bacterial
and fertilization to return cells.
the cells to the diploid state. o The essence of recombinant DNA technology is to remove DNA from
- Bacterial sexual processes cells, manipulate it in a test tube, then put it back into living cells,
are not so regular; they which is mostly done by transformation process.
serve the same aim: to mix o In the case of E. coli, cells are made competent to be transformed by
the genes from two treatment with calcium ions and heat shock. E. coli in this condition
different organisms readily pick up DNA from their surroundings and incorporate it into
together. their genomes.
The three bacterial sexual
processes: Recombination
1. Conjugation: direct transfer mechanisms
of DNA from one bacterial
cell to another. - Best studied in
o The closest yeast, bacteria,
analogue in bacteria to eukaryotic sex. and phage
o The ability to conjugate is conferred by F plasmid [a small circle of - Recombination is
DNA that replicates independently of the chromosome]; bacterial cells mediated by the
that contain an F plasmid are called “F+” and vice versa is “F-“ breakage and
o F+ cells grow special tubes called sex pilli from their bodies; when an joining of DNA
F+ cell bumps into F- cell, the sex pilli hold them together and a copy strands.
Benefits of Recombination 2. Cutting
- Greater variety in offspring generates new combinations of alleles. o Both human DNA and plasmid DNA are cut with the same restriction
- Negative selection can remove deleterious alleles from a population without enzyme.
removing the entire chromosome carrying that allele. o Normally plasmid has only one restriction site while human DNA will
- Essential to the physical process of meiosis, and hence sexual reproduction. have many restriction sites.
o Yeast and Drosophila mutants that block pairing are also defective in 3. Insertion [Ligation]
recombination and vice versa. o Gene is transferred to the new organism.
o It is transferred into the tissue of the organism.
Genetic engineering 4. Transformation
- The artificial manipulation or alteration of genes. o The process of transformation involves the insertion of the
- Also known as Genetic manipulation, Genetic modification, Recombinant desired transgene construct into cells of the recipient plant species;
DNA technology, Gene splicing, Gene cloning scientists isolate tissue or cells from the cultivar they wish to transform
and use one of several methods to insert the transgene into the tissue
Genetic engineering involves: or cells.
o Removing a gene from one organism 5. Expression
o Inserting target gene into DNA of another organism o The result will be a plant with a yield potential close to current hybrids
o Cut and paste process. that expresses the trait encoded by the new transgene.
Restriction Enzyme
Important Terms:
Recombinant DNA: the altered DNA is called recombinant DNA [recombines
after small section of DNA inserted into it].
Genetically Modified Organism: the organism with altered DNA.

Genetic engineering breaks the species barrier:

o Genetic engineering allows DNA from different species to be joined
together.
o This often results in combinations of DNA that would never be
possible in nature; genetic engineering is not a natural process.
o If DNA is transferred from one species to another, the organism that
receives the DNA is said to be transgenic.
o Examples of cross-species transfer of genes:
▪ Human gene inserted into a bacterium
▪ Human gene inserted into another animal
▪ Bacterial gene placed in a plant

Process of Genetic Engineering

1. Isolation
o Removal of human DNA [containing target gene]
o Removal of plasmid [bacterial DNA] from bacterium
Applications of Genetic Engineering
Plants: Weed killer-resistant crops
- Many types of crop plants have bacterial genes added to them.
- These genes make the plants resistant to certain weed killers [bacteria].
- This means that weed killers kill the weeds but do not affect the transgenic
plants.
Animals
- There is a growing trend to
experiment with inserting
human genes into the DNA of
other mammals; transgenic
animals are formed in this
way will produce a human
protein and secrete it into
their milk or even into their
eggs.
Sheep produce human clotting
factor
- A human gene has been
inserted into the DNA of
sheep.
- This allows the adult sheep to
produce a clotting chemical
needed by hemophiliacs to
clot their blood - produced in
the milk of the sheep.
Pharming: production of pharmaceuticals by genetically modified animals [i.e.
sheep, cows, goats etc.]
Microorganisms: Bacteria make insulin
- The human insulin gene has been inserted into a bacterium [E. coli]
- This allows the bacterium to produce insulin for use by diabetics.