Pre Lecture - Molecular Methods Flashcards
What is sensory signaling?
what molecular biology can tell us about how we detect and encode sensory stimuli (pain, touch, taste, smell)
What is the development of the nervous system?
now the brain circuits are formed (axon pathfinding, synapse maturation)
What do genes consist of?
- a promoter region
- a coding sequence that also contains non coding introns
the coding sequence is transcribed into mRNA which is translated into protein
What is the central dogma of biology?
DNA -> RNA -> proteins
By knowing the sequence of the mRNA what can we deduce?
the protein sequence
What can a promoter sequence drive and how can we use this?
drives expression of a gene in specific cell types and we can use it to drive the expression of foreign genes such as GFP
What is gene cloning?
process in which a gene of interest is located and copied or cloned out of DNA extracted from an organism
What do cells that express a gene do?
transcribe the associated mRNA
In what two ways can we measure the mRNA distribution?
- in situ hybridization
- northern blot
What doe insitu hybridization involve?
adding an antisense probe that binds to the mRNA and which contains a label DIG that can be detected by an antibody which is anti-DIG
-antibody is conjugated to an enzyme such that when a substrate is added a blue reaction product is generated
In what two ways can protein hybridization be expressed?
immunocytochemistry
western blot
What does immunocytochemistry involve?
adding an antibody that is directed against the protein of interest ton tissue and cells that are fixed and permeabilized and this antibody is detected via a secondary antibody that is fluorescently labeled
What is a transgenic animal?
animals that are modified by injecting DNA into the early embryo where it is taken up and incorporated into their genome
How is a transgenic animal made?
- link the promoter region to GFP and this can then be used to identify cells that express a gene or direct protein expression to a particular cell type
- link the coding sequence to GFP in frame to generate a fusion protein this can then be used to tell here a protein is expressed which part of the cell
-can be put into a cell under a promoter and fills the cell to identify cell type
-can tether GFP to protein so you can see where the protein is localized in
the cell so protein location determination
What was seen with GFP in transgenic c. elegans?
-can tether GFP to protein so you can see where the protein is localized in
the cell so protein location determination
What is GFP?
protein originally isolated from the jellyfish Victoria aequoria
-absorbs blue light and meets green light
What can we do to know what a protein does?
- introduce the gene into a cell that does not normally express this gene and then can the study the function with patch clamp recording or calcium imaging we can also mutate the gene to identify the functionally important parts of the protein
- KO the gene and see what function in the organisms are perturbed
How do we see and distinguish colors of the rainbow?
we are trichromatic red green ad blue which have a combination and generate colors of the rainbow
What does it mean to be color blind and why is this so common
colorblind people are dichromatic and they only have two pigments to see all colors of the rainbow
Color Blindness Important Info and Experiment
-ask subjects to arrange the tiles in the color of the hue and different types of color blindness exist and they arranged the tile in different ways and you can determine what color blind they are such as protanopia, deuternopia, and tritanopia - red green color blind you can see bluwe but red and green look the same so they seem yellow - X linked chromosome is color blindness
What is trichromacy?
any perceived color can be generated from a combination of three color primaries RGB for additive colors of different wavelength red- long, green - medium, blue - short
What did the search for the GPCR color opsins entail?
-Jeremy Nathans cloned the genes that encode the 3 color opsins and this revealed that
1. the s opsin is very different from the L and M opsins which are very similar to each other (differ in only a few amino acid sequences)
2. the L and M opsins are on the X chrome and are very close to each other
How does color blindness result and what does this mean?
-gene rearrangements that a rose from unequal homologous recombination during meiosis - this means there is not just one type of red-green color blindness and there is not just one type of normal vision
-this means when the chromosomes line up they have to line up right which causes them to sometimes line up with the wrong one like red with green instead of red with red which means when crossing over x occurs one chromosome might just have
green and another could have red red and
green
