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93 Cards in this Set
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What are the features of Bacterial Genome? (4) - single "" and "" (not every bacteria - ex: 2 circular or linear chromosome) - Plasmid .... (how many copies?) - All "" Bacteriophage.... |
What are the features of Bacterial Genome? - single chromosome and plasmids (not every bacteria - ex: 2 circular or linear chromosome) - Plasmid copy number is regulated by the plasmid itself (how many copies?) - All Haploid -Bacteriophage DNA may also be present |
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(Genome) The replicon is the... |
(Genome) The replicon is the collection of nucleic acids which are replicated into the future. |
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(Plasmid) What are features of Plasmids?
- Extra chromosomal... - replicate.... - enable "" ** - #1 feature is.... - cannot have "", "", or "". |
(Plasmid) What are features of Plasmids? - Extra chromosomal genetic elements that replicate autonomously in the cell - replicate as well during cell division - enable special features** - #1 feature is antibiotic resistance - cannot have houskeeping genes, main polymerases, or ribosomal genes. |
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"" grew from microbiology in the ""-"" |
Microbial Genetics grew from microbiology in the 1940s-1950s. |
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Microbial genetics led to... |
Microbial genetics led to molecular biology. |
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Microbial Genetics required the development of... Such as |
Microbial Genetics required the development of model systems for genetic investigations. Such as E.coli and Salmonella typhimurium. |
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(Bacterial Genetics) Early researchers of Bacterial Genetics... (ex: pathogens) |
Early researchers of Bacterial Genetics focused on microbes of practical importance. (Ex:pathogens) |
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Bacterial genetics researchers today also focus on... |
Bacterial genetics researchers today also focus on a greater understanding of the genetic potential of microbes. |
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Why are bacteria ideal genetic research candidates (3) |
1) Bacteria have only one chromosome which allows for easy detection of mutations 2) in early studies, nutritional mutants were used. 3) The capability of studying one gene based on its inability to use or produce a particular nutrient. |
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(Genetic Terminology) What are the features of a Wild type strain: (3) |
(Genetic Terminology) What are the features of a Wild type strain: (3) - strain most like that found in nature - original isolate - source for deriving mutants |
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What are the features of a Mutation? (2) - A Mutation is a .... - Mutations are... |
What are the features of a Mutation? (2) - A Mutation is a change in a gene that disrupts/alters function - Mutations are not always bad |
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The Mutant bacterial cell strain is the... |
The Mutant bacterial cell strain is the strain carrying a mutation relative to the wild type. |
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An allele is defined as a.... Resulting from either a ...,..., or... Of function. |
An allele is defined as a variant of a gene Resulting from either a gain, loss, or change of function. |
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An Autotroph is a.... And often results from a... |
An Autotroph is a mutant that is unable to make a particular compound. Often a mutation in amino acid biosynthesis. (also nucleic acids or other substances) |
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A Prototroph is a... And is often the... |
A Prototroph is a nutritionally wild type strain that does not need any additional growth supplement And is often the auxotrophs parental strain. |
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Genotype is a description of... And generally reflects differences.... |
Genotype is a description of alleles of a given set of genes within an organism And generally reflects differences from wildtypes. |
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Describe Genotype (mutation) using the hisC gene: hisC - .... hisC- : .... |
Describe Genotype (mutation) using the hisC gene: hisC - gene involved in histidine synthesis hisC- : carries a loss of function mutation in hisC gene; genetically wildtype for every other gene |
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Phenotype is the... |
Phenotype is the observable properties of a strain. (Example: His - strain is unable to grow in absence of histidine) |
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(Studying Mutants) Microbial geneticists compare... With a goal of... |
(Studying Mutants) Microbial geneticists compare wild type strains and mutant strains of bacteria With a goal of identifying different alleles of genes. |
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Changes in genes of bacteria are often visible by.. |
Changes in genes of bacteria are often visible by changes in phenotype or growth patterns. |
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(Studying Mutants) What are the features of Selection? (4)
- isolation of.... - Examples: can select for His+... - cannot directly ... - "" |
(Studying Mutants) What are the features of Selection? (4) - isolation of cells with a particular genotype on the basis of growth. - Examples: can select for His+ on basis of growth - cannot directly select for His - - Fast |
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(Studying Mutants) What are the features of Screening? - Identification... - "", "", "no growth" - CAN identify ... - "" |
(Studying Mutants) What are the features of Screening? - Identification of cells based on phenotype. - Color, morphology, "no growth" - CAN identify His- by screening - Slow |
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(Selection) Selectable mutations generally grant... Thus it is useful in genetic research. (ex: ...) |
(Selection) Selectable mutations generally grant a growth advantage under specific conditions. Thus it is useful in genetic research. (ex: conditions that kill wild-type cells) |
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(Selection) Nonselectable mutants confer.... And detection requires... |
(Selection) Nonselectable mutants confer neither advantages nor disadvantages And detection requires screening of a large number of colonies. |
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What are the features of Phenotypic Selection? - Uses a.....the desired gene(s) - "" is commonly used - "" are created and one....
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What are the features of Phenotypic Selection? - Uses a growth medium that will inhibit microbes lacking the desired gene(s) - Antibiotic selection is commonly used - Duplicate plates are created and one lacks a particular nutrient. |
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Screening is more tedious than... |
Screening is more tedious than selection. (with selection, the cells do all the work) |
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Screening can be facilitated with...(identification of "") |
Screening can be facilitated with replica plating. (identification of auxotrophs) |
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Phenotypic Screening can be useful when a... , but doesn't |
Phenotypic Screening can be useful when a mutation has occurred where a colony grows on the full support plate, but doesn't on the partial support plate.
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(Phenotype Screening) Replica Plating can be used.... (uses velvet) |
(Phenotype Screening) Replica Plating can be used to create duplicate plates (uses velvet) |
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What are the features of Patching? (2)
- Patching involves.... - Patching is usually "" and reproducible than..... |
What are the features of Patching? (2) - Patching involves transferring colonies to a gridded plate. - Patching is usually more accurate and reproducible than standard velvet replica plating. |
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What are the features of Mutations? (3) - Generally, DNA replication occurs..., but... - Mutations aren't ...(evolution) - Mutations alter..., sometimes resulting... |
What are the features of Mutations? (3) - Generally, DNA replication occurs without error, but mistakes can occur which result in mutations. - Mutations aren't always bad (evolution) - Mutations alter coding portion of a gene, sometimes resulting in abnormal proteins |
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(Type of Mutation) Silent mutation results... ; usually in the... (no change in phenotype, only genotype)
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(Types of Mutation) Silent mutation results in no change in amino acid sequence of the protein. Usually in the third position of a codon. (no change in phenotype, only genotype) |
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(Types of Mutations) Missense Mutation is a change in... |
(Types of Mutations) Missense Mutation is a change in a codon that results in coding for a different amino acid in the eventual protein. (may affect protein)
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(Types of Mutations) Nonsense Mutation is a change that... |
(Types of Mutations) Nonsense Mutation is a change that forms a stop codon where one should not be found. (bad - strange looking protein)
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(Types of Mutations) Frameshift Mutation is the result of ..., changing how a ribosome...
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(Types of Mutations) Frameshift Mutation is the result of insertions or deletions of nucleotides, changing how a ribosome reads an mRNA molecules and can alter amino acid sequence of proteins. |
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All different types of mutation (silent, missense, nonsense, and frameshift) occur.... |
All different types of mutation (silent, missense, nonsense, and frameshift) occur because of DNA polymerase activity. |
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Reversion is an event when... |
Reversion is an event when a mutation corrects a metabolic abnormality back to the wild-type form. |
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Reversion can be problematic when trying to determine... or....
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Reversion can be problematic when trying to determine mutation rates of a chemical or DNA exchange rates between two microbes.
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(Reversion) The use of "" and "" auxotroph mutant strains decreases....
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(Reversion) Double and Triple auxotroph mutant strains decreases the possibility of a spontaneous reversion mutation. |
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(Spontaneous Mutations) Experiments by "" illustrated "" without "" by using "". |
(Spontaneous Mutations) Experiments by Esther Lederberg illustrated spontaneous mutation without selective pressure (antibiotics) by using replica plating. |
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(Spontaneous Mutation) "" and "" showed that "" to phage infection arises in bacteria without "". |
(Spontaneous Mutation) Luria and Delbruck showed that variable resistance to phage infection arises in bacteria without selective pressure.
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(Evolution in a Test Tube) Describe Richard Lenski's work in 1988 which illustrates the evolution of E.col in a period of 75 days, increasing in fitness:
- cultures which were given extended generation time.... (fresh glucose every day) - their ability to grow.... - this provided evidence of .... - Citrate is usually.... - but they ... |
(Evolution in a Test Tube) Describe Richard Lenski's work in 1988 which illustrates the evolution of E.col in a period of 75 days, increasing in fitness: - cultures which were given extended generation time without selective pressure were compared to original culture that did not have these conditions (fresh glucose every day) - their ability to grow in culture was enhanced over time - this provided evidence of both evolution and the ideal nature of microbes for genetic studies. - Citrate is usually unable to be used by E.coli - but they started using around 40,000 generations |
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What are the three features of Restriction Enzymes (RE) ? - cut DNA.... (viral defense) - Recognition sites...(same read forward and backward) - Similar ends.... |
What are the three features of Restriction Enzymes (RE)? - cut DNA at a specific DNA recognition site (viral defense) - Recognition sites are usually palindromic (same read forward and backward) - Similar ends of cut DNA can be paired together and ligated. |
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Restriction enzymes are always paired with "" This often occurs in ... and is referred to as "".
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Restriction enzymes are always paired with Modification Enzymes. This often occurs in single operon and is often referred to as R/M system.
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Modification enzymes recognize ..... as the.....
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Modification enzymes recognize the same site as the paired restriction enzyme. They add methyl groups to recognition site and prevent Restriction enzyme from cutting them. |
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(Modification Enzyme Example) "" protects DNA from "".They add methyl groups to "" and prevent...
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(Modification Enzyme Example) Methyltransferase activity protects DNA from endonuclease activity (Restriction Enzyme). They add methyl groups to recognition site and prevent Restriction enzyme from cutting them. |
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(Cloning Vectors) Restriction Enzymes allow researchers to stitch.... |
(Cloning Vectors) Restriction Enzymes allow researchers to stitch together fragments of useful DNA into recombinant molecules. |
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(Cloning Vectors) Recombinant molecules (plasmids)can be used to ... |
(Cloning Vectors) Recombinant molecules (plasmids) can be used to clone a bacterial gene of interest. |
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"" are used to insert recombinant DNA molecule into a "" . Three different types of vectors are "", "", or "". |
Vectors are used to insert recombinant DNA into a recipient host bacterial cell. Three types of vectors are plasmids, phages, or cosmids. |
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Plasmid Vector Cloning (pSC101) was first used in "" by "" at "" |
Plasmid Vector Cloning (pSC101) was first used in 1970s by Cohen at Stanford. |
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Cohen cut fragments from.... The transformed strain exhibited....
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Cohen cut fragments from two plasmids carrying antibiotic resistance genes with the same Restriction Enzyme. The transformed strain exhibited traits from both plasmids.
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What are the desirable traits of a cloning plasmid for easier gene cloning ? (6) |
1) origin of replication
2) selectable marker gene (example: amphicilin resistance)
3) multiple cloning site (grouped close together)
4) small size
5) high copy number (easy to extract)
6) plasmid must be closed (circular) |
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Explain the process of Cloning a Plasmid Vector (3 steps) 1 - Cleave....(Recognition site) 2- Mix cleaved ..... (can form different products - different genes come together) 3 - Transform... |
Explain the process of Cloning a Plasmid Vector (3 steps) 1 - Cleave vector and DNA with RE (Recognition site) 2- Mix cleaved vector fragment and genomic DNA fragments and ligate. (can form different products - different genes come together) 3 - Transform ligation products into E.coli cells and spread onto plate... |
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(Screening for Transformed Cells) The x-gal system can be used to detect transformed cells such that... |
(Screening for Transformed Cells) The x-gal system can be used to detect transformed cells such that the transformed cells will have a blue color and the rest of the colony will be white. |
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What are the features of X-gal screening: - When new..., "" is affected. - Produces... (plasmid present) - X-gal reacts.... - if no plasmid, .... |
What are the features of X-gal screening: - When new gene is added, Lac Z is affected- Produces abnormal B-galactosidase (plasmid present) - X-gal reacts with abnormal B-galactosidase - if no plasmid, functional B-galactosidase would form |
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(Shuttle-vector Plasmids) certain host cell types may be ...
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(Shuttle-vector Plasmids) Certain host cell types may be restricted by the origin of replication. |
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Shuttle Vector Plasmids have multiple.... This expands the range of... |
Shuttle-vector plasmids have multiple types of origins of replication. This expands the range of host cell types the plasmids can be inserted into. |
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A Phage vector results when.... Then, this phage vector is.... |
A Phage vector results when a viral DNA is mixed with the fragment of interest which occurs when RE separate the genes requires for lysogenic pathway and converts the pathogen into a lytic virus. Then this phage vector is added to the host cell. |
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(Phage vector example) "" can carry "" sized fragments.
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(Phage vector example) Lysogenic lambda phage can carry ~20 kb fragments. |
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Cosmids are.... such that only the critical phage "" (packages genome..) ....
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Cosmids are phage genomes that exclude nearly all phage DNA (leaves more room for fragment) such that only the critical phage "cos" packaging recognition site (packages genome into virus) remains.
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Other elements of Cosmids include a "" and an "". Cosmids have "" and can replicate... |
Other elements of a Cosmids includes a multiple cloning site and an antibiotic selection marker. Cosmids have oriV and can replicate autonomously, thus don't change virus lytic/lysogenic pathway. |
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Cosmids can typically carry "" - "" kb fragments but must be about... |
Cosmids can typically carry 35-45 kb fragments but must be about 50 kb in total for successful implementation into the viral genome head (fragments can be very large for cloning) |
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What are the features of Transformation? - introduction of.... - Doesnt require... - some bacteria are... - Other bacteria can be artificially... |
What are the features of Transformation? - introduction of extracellular DNA directly into an organism - Doesnt require cell-to-cell contact- some bacteria are naturally competent for transformation - Other bacteria can be artificially induced to become competent for transformation |
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What are two ways to induce transformation into a cell? - Chemically competent cells... - Electro-competent cells... |
What are two ways to induce transformation into a cell? - Chemically competent cells are treated with calcium cations. - Electro-competent cells are treated with electroporation. |
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Describe the steps of Transformation in naturally competent cells 1 - ssDNA... 2 - coated... 3 - ssDNA identifies.. and introduced... |
Describe the steps of Transformation in naturally competent cells 1 - ssDNA is sucked up by cell 2 - coated with RecA protein 3 - ssDNA identifies region of homology (similarity) and introduced to organisms chromosome by homologous recombination |
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In natural bacterial transformation why is the DNA single-stranded and what happens if homologous recombination doesn't occur? - Single-stranded so it is... - If homologous recombination doesn't occur then.... |
In natural bacterial transformation why is the DNA single-stranded and what happens if homologous recombination doesn't occur? - Single-stranded so it is not susceptible to attack by Restriction enzymes. - If homologous recombination doesn't occur then ssDNa will be lost and degraded as a nutrient. |
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Conjugation is the transfer of .... |
Conjugation is the transfer of DNA from cell to cell via direct contact/sex pilus formation. |
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What are the features of the Conjugation mechanism (5) - F plasmid |
1) F plasmid carries gene to form sex pilus "bridge" between 2 cells. (tra genes)
2) F plasmid can be copied and sent across the sex pilus bridge (oriT transfers first) into a recipient cell.
3) F plasmid turns an F- cell into an F+ cell capable of conjugating with another F- cell. (end of process both cells will be F cells)
4) Tra genes are essential for mediating the conjugation process
5) OriT is also required such that the arrow shows direction of plasmid through sex pilus bridge (genes that follow include oriV)
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Describe the steps of Conjugation (bacterial sex) 1 - Sex.... 2 - After connection, ....(tra genes) 3 - 1 strand of..., then replication ....resulting in .... 4 - results in... Tra genes also include.... 3 or more cells.... |
Describe the steps of Conjugation (bacterial sex) 1 - Sex pili pull cells together 2 - After connection, form sex bridge (tra genes) 3 - 1 strand of F plasmid transfers to recipient, then replication occurs in both cells resulting in 2 double stranded DNA molecules. 4 - results in 2 F+ bacteria Tra genes also include recognition genes to recognize F- cells 3 or more cells can do conjugation all at one |
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(Conjugation) The F plasmid can integrate into .... by ... |
(Conjugation) The F plasmid can integrate into the host chromosome by homologous recombination. (Produces the Hfr plasmids) |
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(Conjugation) The F plasmid is an "" ( ... Definition)
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(Conjugation) The F plasmid is an episome (DNA that can integrate into the chromosome or exist autonomously). |
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(Conjugation) The F plasmid creates new opportunities to...
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(Conjugation) The F plasmid creates new opportunities to pass genetic information from one cell to another. |
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What is a Hfr (high frequency of recombination) bacterial strain produced through conjugation?
An Hfr strain is a bacteria with ...into its "" through ""
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What is a Hfr (high frequency of recombination) bacterial strain produced through conjugation?
An Hfr strain is a bacteria with a conjugative plasmid (F plasmid) integrated into its chromosomal DNA through homologous recombination |
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Hfr bacterial strain DNA transfer can be used to.... |
Hfr bacterial strain DNA transfer can be used to "map" the location of genes in the host chromosome. |
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Briefly describe how Hfr DNA strain transfer can be used to map alleles passed between each cell 1 - place Hfr.... 2- Allow ...(requires "" and "" conditions) 3 - Interrupt.... 4 - New genes within.... |
Briefly describe how Hfr DNA strain transfer can be used to map alleles passed between each cell 1 - place Hfr with F- cell 2- Allow conjugation to occur (requires stable and gentle conditions) 3 - Interrupt mating to disrupt mating bridge 4 - New genes within F- cell will do homologous recombination with original genome. |
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F' plasmids result when an....
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F' plasmids result when an incorporated F plasmid excises itself by homologous recombination. |
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What are the features of F' plasmid generation? (2)
- Excision is "" and some... - When the F' plasmid conjugates,... |
What are the features of F' plasmid generation? (2) - Excision is inaccurate and some host cell DNA is excised as well. - When the F' plasmid conjugates, it sends the host cell DNA to the recipient. |
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An example of F' plasmid generation would be Hfr E.coli cells excises itself and becomes E.coli genome + F' plasmid (contains some of E.coli' chromosomal DNA) |
An example of F' plasmid generation would be Hfr E.coli cells excises itself and becomes E.coli genome + F' plasmid (contains some of E.coli' chromosomal DNA) |
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(Uses of Conjugation) Triparental conjugation is the event when conjugation occurs using... |
(Uses of Conjugation) Triparental conjugation is the event when conjugation occurs using recombinant plasmid (only OriT) lacking the required tra gene and a helper plasmid with the tra gene. (OriT and Tra genes) |
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(Uses of Conjugation) Triparental Conjugation is useful because there is more room... |
(Uses of Conjugation) Triparental conjugation is useful because there is more room in the recombinant plasmid for a desired DNA fragment. |
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What are the features of Triparental conjugation: (4) - Helper strain contains... - Recombinant DNA.... (not F plasmid) - Helper strain will conjugate with.... - Requires 3 plasmids which are.... |
What are the features of Triparental conjugation: - Helper strain contains a F plasmid which has tra genes and an oriT - Recombinant DNA doesnt have tra genes (not F plasmid - Helper strain will conjugate with donor strain by encoding the proteins required for transfer. - Requires 3 plasmids which are helper strain, donor strain, and recipient strain. |
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What are the features of Transposition (jumping genes)? - Transposition is the...
- Transposable elements can move....(transposons) -Transposition was first discovered... (corn) |
What are the features of Transposition (jumping genes)? - Transposition is the movement of DNA via mobile genetic elements. - Transposable elements can move within and between genomes. (transposons) -Transposition was first discovered by Barbara McClintock (corn) |
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Transposition can be subdivided into:
Insertion sequences only...
Transposons contain other genes.... |
Transposition can be subdivided into:
Insertion sequences only encode the proteins needed for transposition
Transposons contain other genes in addition to those needed for transposition |
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Mechanisms of Transposition requires "" (both) and "" (only "") |
Mechanisms of Transposition requires transposase (both) and resolvase genes (only replicative transposition) |
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What are the two types of Transposition? Replicative transposition copies... (Has IR sequences and requires "") Non-replicative transposition.... ( had IR sequences) |
What are the two types of Transposition? Replicative transposition copies the elements (genes) and moves the copy to another location (Has IR sequences and requires Res genes) Non-replicative transposition cuts and pastes the elements into a new location ( had IR sequences) |
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A simple way to identify transposon genes in addition to them having.... is that it.... |
A simple way to identify transposon genes in addition to them having IR genes on both sides is that it will have a few extra bits of Target DNA sequence on either sides. |
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The frequency of transposon movement is "" and the "" and....is random! |
The frequency of transposon movement is low and the target sequence and location of transposon landing is random! |
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Transposition can be used to.... (effect way to "") |
Transposition can be used to disrupt functional genes and observe phenotypic genes (effect way to generate mutants) |
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Describe how a Suicide vector plasmid carrying a transposon creates a mutant 1 - recipient cell...(with transposon) but... 2- transposition... 3- ....for desired disruption (mutation) (selection is possible since....) |
Describe how a Suicide vector plasmid carrying a transposon creates a mutant 1 - recipient cell gets the plasmid (with transposon) but the plasmid cant replicate 2- transposition can occur at random 3- Screening and/or selection for desired disruption (mutation) (selection is possible since antibiotic marker is present) |
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What are the features of Viral Transduction through Transposition - Lytic Virus accidently.... - Lytic Virus delivers that .... - Lytic Virus is usually unable... - ""must occur - "" (virus contains "") |
What are the features of Viral Transduction through Transposition? (5 steps)
- Lytic Virus accidently packages a fragment of host cell DNA - Lytic Virus delivers that fragment instead of viral DNA to next cell - Lytic Virus is usually unable to replicate because it lacks the viral genome - homologous recombination must occur - Low frequency event (virus contains donors DNA) |
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Historically, co-tranduction frequency was used to map bacterial genomes such that genes that were closer to a known "marker" gene (lytic virus genome) would be transduced with that marker more frequently than the ones farther away. |
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What are the features of Co-transduction (uses lysogenic virus)?
- can be used to modify bacteria - has only been performed in vitro so far |
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What is an example of Co-transduction (uses lysogenic virus)
experimental modification of a common vaginal tract microbe to express and secrete a chemokine with anti-HIV activity (given through Lysogenic virus) |
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