Lecture 1- Microbial Ecology/Communities Flashcards
give some examples of culture independent analyses
gen staining methods, flurorescent in situ hybridisation (FISH), PCR methods of microbial community analysis, environmental genomics and related methods
write a quick note on DAPI
[4’,6-diamindino-2-phenylindole] is a general fluorescent stain. It binds strong;y to adenine-thymine rich regions in DNA and hence stains DNA. It is used for organism identification in natural samples. Some stains can differentiate between live and dead cells, and so fluorescent antibodies that are specific for one strain/small group of related cells can be used.
What kit can be used for microscopy and quantitative assays
the LIVE/DEAD BacLight Bacterial Viability Kit. Compromised membranes (dead) = stained RED and intact membrane (alive) = GREEN
write a note on Live Bacterial Gram Stain Kit
Fluorescence staining kit for live cell gram stains. the dye binds to N-acetylglucosamine in peptidoglycan, and it stains G pos cells with RED fluorescence.
The kit has DAPI too- to stain DNA blue. Gpos cells will be stained red on the outside and blue in the middle, and G neg cells will be all blue.
how does viability PCR work
these dyes are membrane impermeable, meaning only dead cells can uptake them. the dyes become reactive and covalently attach to DNA. the modification of the the DNA prevents amplification of next PCR reactions.
this is good for selectively detecting live bacteria via PCR
What does GFP do?
green fluorescent protein makes cells autofluorescent and can help track cells introduced to an environment
define phylogenetics
analysis of molecular sequencing data in order to study evolutionary relationships between organisms
define phylogeny
the evolutionary history of a group of organisms
how do we get phylogenetic information
from nucleotide sequence data
why are rRNA genes used for phylogenetical analysis
proposed by Carl Woese- they are universally distributed, funtionally constant and are sufficiently conserved as they are slow changing
which rRNA subunit is used for prokaryotes and eukaryotes
16S in prokaryotes
18S in eukaryotes
