Term
| Hypothesis for typing systems? |
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Definition
| Series of isolate obtained from an epidemiological cluster are clonally related. |
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Term
| Characteristic of typing systems |
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Definition
| It is only useful if there is stability within a strain of organism(identify same organisms) and diversity within a species (differentiate between different organisms within a same species) |
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Term
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Definition
based on presence or absence of metabolic/biologic activities.
* organsims can alter expression of biologic properties(mutation, environment) so its not as reliable as molecular methods. |
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Term
| Genotypic systems(molecular) uses what as the template? Why? |
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Definition
| nucleic acids. Diversity among microbial species. Can show evolutionary divergence. |
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Term
| Advantages of using nucleic acid as a template for ID? |
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Definition
Relatively Stable
Ubiquitous in nature (found in all organisms).
Purification is identical for all organisms. |
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Term
| PCR - what is it? Advantages? |
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Definition
Target site amplification.
Rapidly and exponentially amplify specific DNA sequence.
Rapid
Sensitive
Specific. |
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Term
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Definition
| Need a template DNA, 2 primers to replicate DNA (one for each strand, 10-20 bases long), free base pairs, DNA polymerase(TAQ - thermostable, because of the use of heat in the procedure). |
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Term
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Definition
3 cycles:
Heat is up - to denature it, turns into two single strands of DNA.
Cool to annealing temperature - binds primers to complementary DNA sequences.
Raise temp to 72 - DNA polymerase can start to build new DNA. |
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Term
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Definition
| Separates amplicon - based on size and charge. DNA is negatively charged due to phosphate backbone. Travels to anode (+). |
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Term
| Visualization of Separated DNA - |
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Definition
| Add a die that attaches to DNA and fluoresces in UV light. ex. Ethidium Bromide. |
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Term
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Definition
| Uses a computer analyses as it processes. No electrophoresis needed, takes about 10 mins. |
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Term
| Pulse Field Gel Electrophoresis |
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Definition
High molecular weight DNA, is cut with restriction endonuclease. Electrophoresis is done systematically with different angles and currents.
Used for epidemiological purposes.
HEXAGON. |
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Term
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Definition
| Highly discriminatory - differentiates between strains of the same species. Out break invesigation. Track evolution of organisms. |
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Term
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Definition
| Equipment is expensive, requires skilled technologist. |
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Term
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Definition
Indistinguishable - banding pattern is the same.
Closely Related - 2-3 band differences. Same bug, mutation occurred.
Beyond six - not related. |
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Term
| 16s rRNA gene sequencing - premis. |
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Definition
rRNA common to all bacteria.
Based on the premis that all bacteria contain 16s ribosomal RNA gene. Its an essential gene.
This gene evolves very slowly. Molecular clock – track changes in genes over time using this gene. |
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Term
| Two different areas of the 16s rDNA sequencing |
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Definition
Conserved region - detects bacteria.
Divergent - variable within a species. Help to identify the bacteria.
Design a primer to target the conserved region. Once you have that amplified, sequence the variable regions which are different from bacteria to bacteria. |
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Term
| How do you identify bactria using 16s rRNA gene sequencing? |
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Definition
| PCR primer designed to anneal to conserved regions. Can sequence the variable region and input data into a data base to find what it is analogous to. |
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Term
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Definition
No need for viable organisms. Can be dead bacteria.
Detection and ID of organisms in people who have already started Tx.
Detection of slow/non growing organisms. |
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Term
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Definition
Only useful from normally sterile sites - contamination is a big problem because it is so sensitive.
Only useful for monomicrobial infections.
Slow process. |
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