Microbial genetics Flashcards
Summary Basic concepts Prokaryotes vs Eukaryotes Mutants production Exchange of genetic material Restriction enzymes CRISPR overview
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Basic concepts
DNA
deoxyribonucleic acid, the genetic material of cells and some viruses.
Basic concepts
RNA
ribonucleic acid, functions in protein synthesis as messenger RNA, transfer RNA, and ribosomal RNA.
Basic concepts
Gene:
is a sequence of nucleotides in DNA that codes for a functional product.
Basic concepts
Chromosome:
a genetic element carrying genes essential to cellular function.
Prokaryotes typically have a single chromosome consisting of a circular DNA molecule.
Eukaryotes typically have several chromosomes, each containing a linear DNA molecule.
A chromosome consist of an association of genes and some protein. The chromosome is about 1mm long, which is about 1000 times longer than the typical bacterial cell. Therefore, it is looped, folded and packed tightly inside the cell.
Basic concepts
Promoter:
DNA sequence immediately in front of a gene that is needed to activate the gene’s expression.
Basic concepts
A gene can:
- Stand-alone, operating independently from other genes.
- Exist in tandem with other genes, in an operon. All genes in an operon are situated head to tail on the chromosome and are controlled by a single regulatory sequence located in front of the first gene.
Basic concepts
Plasmid:
an extrachromosomal genetic element that is not essential for growth and has no extracellular form.
Basic concepts
Proteins:
are macromolecules that play many functions in the cell consisting of one or more polypeptides. They are used for support, storage, transport of other substances, defense against invaders, and catalytic enzymes. Proteins are composed of repeating units called amino acids
Prokaryotes Vs Eukaryotes
- Genome Size
- Noncoding DNA
- Splicing
- Genome in
- Regulatory pathways
- Cell division
- Genetic material exchange
- 0.5 to 9.4 Mbp
- 9 to 100,000 Mbp
- No (rare)
Yes - No
Yes - Nucleoid
Nucleus - Simple
Complex - Binary
Mitosis - Horizontal transfer
Meiosis
Mutant Production
Mutation:
An inheritable change in the base sequence of the genome of an organism
Point mutation
2 points
- Substitution, is a single point mutation that occurs when a single nucleotide is replaced with a different nucleotide.
- A point mutation results in a base pair substitution after replication and possibly a mutant protein after transcription and translation
Mutation Rate
- The probability that a gene will mutate when a cell divides
- Spontaneous mutation rate for the average gene is 0.000000001.
- The presence of a mutagen increases the rate of mutation to 0.00001 to 0.001.
Mutagen
Chemical, physical or biological agents that can increase rate of mutation and so induce mutations.
e.g.
- Nitrous acid (chemical)
Deaminates Adenine and Cytosine
Affects DNA complementation
Mutant Isolation
Positive Selection:
growing the culture on a medium that will allow for the growth of only the mutant colonies. E.g., Penicillin resistant mutants grow on a medium that contains penicillin.
Mutant Isolation
Negative Selection:
used to identify mutants that have lost the ability to perform a certain function that their parents had. E.g., Auxotrophic mutants (bacteria that have lost the ability to synthesize an essential nutrient).
Mutant Production
3 points
- Suppressor mutation: is reversal of mutant phenotype by another mutation at a point of DNA distant from that of original mutation.
- All genes are susceptible to mutations, but not all mutations are expressed.
- Lethal mutation is harmful destroy the vital functions.
Exchange of genetic material
Horizontal gene transfer
3 points
- transformation
- transduction
- conjugation
Exchange of genetic material
Horizontal gene transfer
Transduction
5 points
- Use of bacteriophages to transfer DNA between cells
- Genetic material exchanged via viruses. E.g. bacteriophage lambda
- No physical contact
- Promotes drug resistance, e. g. Penicillin resistance in Staphylococcus
- Helps genetic mapping and engineering
Exchange of genetic material
Horizontal gene transfer
Transformation
3 points
- Acquisition of exogenous DNA fragments from the environment. E. g. plasmids
- Antibiotic resistance
- Pesticide resistance in crops. E.g. Agrobacterium tumefaciens
Exchange of genetic material
Horizontal gene transfer
Conjugation
5 points
- Cell to cell contact required
- DNA exchanged via Pili
- Large segments could be transferred
- Transfer is nonspecific
- Drug resistance
Restriction Enzymes
3 points
- Arestriction enzymeis anenzymethat cutsDNAat or near specific recognitionnucleotidesequences known asrestriction sites.
- origin: To protect themselves against invading virus DNA/RNA
- Methylation avoids its own degradation
Restriction Enzymes 3 uses
- To generate a restriction map: provide useful information in characterizing a DNA molecule.
- Restriction Fragment Length Polymorphism (RFLP) is a tool to study variations among individuals & among species.
- Most widely used in recombinant DNA technology.
What is CRISPR?
C Clustered R Regularly I Interspaced S Short P Palindromic R Repeat
two main components of CRISPR-Cas9:
- The Cas9 enzyme: which cuts DNA
2. A guide RNA: that guides these molecular scissors to the sequence we want to cut
Editing a gene using CRISPR/CAS9 technique
4 points
- create genetic sequence called “guide RNA’ that matches piece of DNA want to modify
- add to cell with protein Cas9 which acts like scissors (cuts DNA)
- guide RNA homes in on target DNA sequences, Cas9 cuts it out
- another peice of DNA swapped into place of old DA and enzymes repair cuts.
= DNA edited
