Module 6: Mechanisms of genetic variation and microbial DNA technologies Flashcards
what is screening
process whereby individual cells (or clone of cells that have grown into a colony) are checked one by one to determine if they have the mutant phenotype
- replica plating
what is selecting
subjecting a population of cells to an environment condition that only allows for the growth/survival of the desired mutant phenotype
- a very large number of cells can be tested by the selection and only selected mutants will growth the colony
what is an auxotroph
- unable to grow on a minimal media that the wild type parent can
- can now be used as the parent or starting strain from which to select for revertant strains that have undergone further mutations that afford the cell to further undergo growth in minimal media
what is proof reading
10-100 fold reduction in errors rates while copying the genome
what is a mismatched pair
- next line of defence
- repairs trails behind the advancing replication fork, and scans the newly formed double helix for mispaired bases
what is an excision repair
a repair pathway that detects distortions in the DNA double helix
- it is not specific to any type of error, it is an effective and versitile repair system
- it excises (removes) a short sequence of DNA from one of the strands
what is a base exsision repair
- carried out by a family of enzymes that each individually recognize specific unnatural bases within the DNA double helix
- enzymes are glycosylases, they remove damaged base from the sugar phosphate backbone, leaving an abasic site
what is direct repair
- other enzymes have the ability to recognize specific changes to bases in te duplex and reverse those changes without removing the affected base or region
what is recombinational repair
- when a cell carries more than one copy of the chromosome (which is the case for regions that have been replicated, even haploid bacteria)
what is the SOS response
- is high alert response that coordinates multiple repaire pathways in their responses following a relatively large degree of damage to the cells DNA
- it is regulated by a common repressor protein (LEXa ) which is inactivated when RecA protein is bound to excess single strand DNA arising from stalled replication and/or incomplete DNA repair
what is horizontal gene transfer
transfer of bits of genetic information from a non parent cell is given
- this process is not as frequent as the exchange that occur during sexual reproduction, it is very powerful in that it brings new genetic information and tools from multiple different selection and evolutionary experiences, within the recipient species having had to proceed through the same selection
what is transposable element
- ## small DNA molecule that carries the genes for transposition (hopping to a new location) and can move around between genomes
what is transposons
- more complex transosable elements that carry more than just their recombinase gene
- many bacterial transposons have been found that carry antibiotic
what is recombination
bacteria is characterized by joining DNA from one DNA molecule (the donor) to another DNA molecules (the recipient)
what is homogous recombination
two dna molecules that are very similar in nucleotide sequence. the mechanism depends on a homology search by recombination specific enzymes
- this is also a mechanism of DNA repair in bacteria
what is site specific recombination
- happens between two DNA molecules that do not have extensive areas of homology
- is catalyzed by enzymes called recombinases = transposition occurs
what is conjugation
- bacteria does not undergo sexual reproduction, they have mating system
a number of bacteria plasmids have been identified that have the ability to transfer a copy of their DNA from a donor cell to recipient cell
what is transformation
- the ability of an organism to take up foreign DNA from its environment, and if it shares sufficient homology with its chromosome, recomdining that DNA into its genome
what is competent
- when a cell has turned on its DNA uptake machinery
what is natural compentency
- carry the potentialn to turn on the competent function,
what is transduction
- transfer of genes between bacterial or archaeal cells by viruses
- major source of horizontal gene transfer in nature
what is generalized transduction
- random packaging of host DNA by a virus
- has the ability to randomly transfer any gene from the chromosome
- viruses that facilitate generalized transduction are typically lytic phages that are dedicated to the lifestyle of an infect (replicate, package and lyse (destroy the cell) repeat)
what is specialized transduction
- gene transfer carried out by viruses that are lysogenic phages
what is a restrictive enzyme
cut or break any DNA within the cell that was unmethylated at those same specific DNA sequences
- DNA endonucleases
how do you purify DNA
- amplification (replications of cells carrying only that clone)
- purification of the vector with the insert (creating a solution with many copies of the same DNA)
- cutting that DNA with a specific restrictive enzyme, or combination of restrictive enzymes
- separation of the created fragment based on their size
- collection of the fragment of interst for further analysis
what is gel electrophoresis
separating DNA molecule based on size (length)
- typically agarose or polyacrylamide gel,
- the pores within the Gel allow for movement of the negatively charged DNA molecules towards the positive pole
what is reverse transcriptase (RT)
does the reverse of transcription - it starts with an RNA template and makes a complementary DNA or cDNA copy
what are cloning vectors
-small piece of self replicating DNA that can be stably maintained in an organism, and into which a foreign DNA fragment can be inserted
- plasmid vectors, phage and other viral vectors, cosmids and artificial chromosomes
what is a genomic libraries
- collections of ramdomly - fragmented pieces, each individually ligated into a copy of the cloning vector and trasnformed into the host cell
