BIOL240 Chapter Notes -Lambda Phage, Holliday Junction, Sos Response

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MODULE 6
Part A: An Introduction to the Terminology of Molecular Biology
History
- E. coli and Salmonella developed in first genetic system
Important Terms
- Mutant: strain carrying mutation
- Mutation: change that occurs within gene to make another allele of gene or disrupt
function of gene
- Mutation gain/loss of function, change of function at DNA level
- Genotype: description of all alleles within organism
- Phenotype: observable properties of strain containing mutation
- Wild-type: strain found in nature original isolate of strain that all mutants are derived
from
Part B: The Phenotypes of Mutations Selection vs. Screening
Selectable Phenotype
- Bacteria plated out on growth medium under certain conditions that will allow
phenotype of interest to grow
- Power of genetics ability to use selection vs. screening
- Screenable phenotype cannot be selected but can be differentiated
Isolation of Mutants
- Error occurring during replication leads to evolutionary change
- Mutations arise spontaneously
Replica Plating
- Culture of wild-bacteria subjected to mutagenic conditions to increase mutation rate
- Culture diluted and placed on complex medium tor single colonies on master plate
- Auxotroph: organism that developed nutritional requirement through mutation
- Potential auxotrophs do not grow on minimal medium
Part C: Types of Mutations
Examples
Auxotrophs
- loss of enzyme in biosynthetic pathway
- inability to grow on medium lacking nutrient
Cold-Sensitive
- inactivated protein at low temperature
- inability to grow at temperature normally supporting growth
Drug-Resistant
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- Permeable to drug
- Growth on medium containing growth-inhibitory concentration of drug
Noncapsulated
- Loss or modification of surface capsule
- Small rough colonies and not large smooth ones
Nonmotile
- Loss of/non-functional flagella
- Compact colonies instead of flat, spreading ones
Pigmentless
- Loss of enzme in biosynthetic pathway leading to loss of pigment(s)
- Presence of different/lack of colour
Rough Colony
- loss or change in lippolysaccharide outer layer
- Granular/irregular colonies
Sugar Fermentation
- Loss of enzyme in degradative pathway
- Lack of colour changes on agar with sugar and pH indicator
Temperature Sensitive
- alteration of essential protein so becomes more heat-sensitive
- inability to grow temperature normally supporting growth
Virus Resistant
- loss of virus detector
- growth in presence of large amounts of virus
Shifts in Reading Frame
- deletion or insertion mutation
- To avoid shifs in reading frames add or delete 3 amino acids
Part D: Mutagens
- Mutagens can increase frequency of mutations
The SOS Response
- several genes with expression induced to respond to DNA damage
- Some genes can repair damage
- sometimes DNA damage is so severe cell dies if not repaired
- UvrA protein error-gree repair of DNA damage
- UmuD protein error-prone repair of severe chromosomal damage
- SOS response last resort of the cell
- RecA detects DNA damage lexA (binds operator in front of gene) cleaved
- Single stranded DNA = signal to RecA that DNA damage has occurred
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Part E: The Ames Test
Test for Mutagenicity
- determines whether product or chemical is mutagenic
- if mutagenic, possibly carcinogenic
- Salmonella strain containing point mutation in histidine biosynthetic pathway is used
- Salmonella strain = histidine auxotroph
- measures rate of mutation
Part F: Genetic Recombination
Introduction
- DNA transferred from one bacterial strain to another
- Transformation: transfer of DNA between bacterial strains
- Transduction: involves use of bacterial phage to transfer segments of chromosome
- Plasmids also involved in transfer of genetic material
The Mechanism of Homologous Recombination
- Recombinatoin event allowing DNA molecules to be joined
- Occurs in multicellular eikaryotic organisms
- High level of recombination during meiosis
- Can also occur in bacteria when segments of DNA inserted from one strain to another
- Segments have different alleles exchange of alleles between organisms
- Single stranded donor molecule base pairs with recipient molecule (RecA)
- strand bound to recipient displaced by donor molecule
- Cross-strand exchange develops with formation of Holliday junction
- Splicing of donor and recipient pathces or splices recombination
DNA Transfer
- Donor free DNA liberated out of cell
- Taken up by Recipient cell
- Transformation: transfer of DNA
- Transduction: transfer of host genes from one cell to another by virus
- Conjugation: transfer of genes from one prokaryotic cell to another by cell-to-cell
contact
- Plasmid: extrachromosomal genetic element that is not essential for growth and has no
extracellular form
- Plasmid transferred through conjugation
Detection of Recombination
- selectable markers or phenotypes determine whether recombination has occurred
- ability to select cells out of background selectable phenotype
Transformation and Transduction
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Document Summary

Part a: an introduction to the terminology of molecular biology. History: coli and salmonella developed in first genetic system. Mutation: change that occurs within gene to make another allele of gene or disrupt function of gene. Mutation gain/loss of function, change of function at dna level. Genotype: description of all alleles within organism. Phenotype: observable properties of strain containing mutation. Wild-type: strain found in nature original isolate of strain that all mutants are derived from. Part b: the phenotypes of mutations selection vs. Bacteria plated out on growth medium under certain conditions that will allow phenotype of interest to grow. Power of genetics ability to use selection vs. screening. Screenable phenotype cannot be selected but can be differentiated. Error occurring during replication leads to evolutionary change. Culture of wild-bacteria subjected to mutagenic conditions to increase mutation rate. Culture diluted and placed on complex medium tor single colonies on master plate. Auxotroph: organism that developed nutritional requirement through mutation.

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