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UConn MCB 2210 - Cell Theory

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StudeerSnel wordt niet gesponsord of ondersteund door een hogeschool of universiteitCell Bio Mcb 2210 Class NotesCell Biology (University of Connecticut)StudeerSnel wordt niet gesponsord of ondersteund door een hogeschool of universiteitCell Bio Mcb 2210 Class NotesCell Biology (University of Connecticut)Gedownload door Mark Salib ([email protected])lOMoARcPSD|48571371/20/16Intro lecture Cell Theory- All living things are made of cells- The cell is the basic structural unit of living things- Cells can only arise by division from preexisting cellso Core concepts that are still believed to be true todayBasic prosperities of cells- List is common in most cells but not all cells have all these properties - There are many exceptions within our bodiesCentral Dogma- How the cells exhibit all their properties - Information that allows cells to do and express things are encoded in their genome - Activation  transcription  Processing  Translation - More complex than what diagram shows - Vast number and types of proteins encoded in the genomes of different eukaryotes Cells are dynamic- Not all the info in a cell is encoded directly in DNA in a form we can read- Cells are either methylated or acetylated 1/22/16- All cells have a common ancestry - First eukaryotic ells are more related to archea than bacteria - Prokaryotic cells arose first therefore they’re simpler cells but that doesn’t mean they’re inferior - Single celled organisms are generalists therefore complicated and do many functions Core Tools of Cell BiologyType of cells cell biologists study- Primary cells: cells that are isolate straight from a living organism that can be cultured to study cell specific functions.o Many are differentiated so they aren’t going to divide anymore/hang around in the dish longero Can bring limited values bc we can’t understand their growth factors that keep them around longer. o Cells right out of the organ. Not many are differentiated so they don’t divide anymore- Transformed cells: cells that have become cancerous because they divide continuously therefore easy to maintain in lab- As technology increases, we can study cells in vivo/in their real environment instead of pulling them outin a dish bc you don’t know if how they act in a dish is relevant in their lives. - Light microscopy: one of the biggest toolso Limited (around 200 nm) in contrast, magnification and resolving power, limiting what can be detected with themo The quality of microscopes have been improving but again, there are hard limitations o Cells are mostly water therefore they’re very transparent. Light passes through them bc they have poor contrast. Need to enhance their contrast (poor contrast due to water)- Transmitted light microscopy: magnify a specimen using one or more lenseso White light passes thru the specimen before being collectedo Cells are neither reflected or absorb much light so contrast is poor and little details can be madeo Contrast is the difference in intensity between an object and its backgroundo Bring out detail, we need to exploit changes in the phase of light or stain the object to make it darker o Phase contrast or differential interference contrast  techniques for enhancing contrast - Magnification vs. resolution Gedownload door Mark Salib ([email protected])lOMoARcPSD|4857137- Magnification is how much you blow up on image. Depends on lens.- Resolution= how far apart two objects have to be to be seen as 2 different objects- Depends on wavelength. Shorter wavelength = better resolution as well as how good ur lens are. Numerical aperture (how well lens can gather cone of light coming out of the specimen)- Resolution of conventional light microscopes is ½ the wavelength of light being used 1/25/16- Ability to resolve doesn’t mean ability to detect. - Just means you won’t be able to see if it’s smaller than a certain nm- Anything smaller will look the default nm- Epifluorescence microscopy improves contrast and allows specific structures to be labeled via staining. - Smaller wavelength = higher energy - Illuminates molecules w/ fluorophores via higher energy light- Antibodies: immune proteins that bind to specific proteins.o To make, obtain protein, pure of interest and inject it to animal. o Monoclonal vs. polyclonal antibodies - Immunocytochemistry only works for nonliving cells- Western Blotting- used to detect proteins with antibody detection. o Smallest proteins can elude through SDS PAGE gel fastero Gel separates protein by size of moleculeo Use electric field to separate protein- Fluorescent proteins can keep the cell in tack so we never kill the cell - To get plasmid across membrane is called transfection o Infect cells with foreign DNA to cause a desired expressiono Transfection can be transient or stable (expression from plasmid or DNA integrates into genome/heritable)o Transgenic lines of animals can be generated by stable transfection of germ cellso We can transfect mutant molecules to be active all the time by activating a protein to turn it ono Mutant molecules that are dominant negative don’t function right and block the function of the cell’s own version of the molecule  Mutant form of enzyme can’t catalyze enzymatic reaction aka doesn’t work/non-functional therefore causes the mutant form of the cell to not work as well. Has ability to even block function of normal enzymes1/27/16- A multi-color fluorescent image uses different proteins and antibodies as well as DNA dyes to add color to a black and white microscopic image- 200 nm is the lowest resolution of light microscopy you can see so any object smaller than 200 nm will always appear 200 nm, regardless of it’s size- siRNA help transfect things into cells using new technologies that exploit RNAi mechanisms to allow usto knock out specific proteins of interest in cells by targeting mRNAso **Don’t memorize image on this slide. Not a genetics class. Just understand generally what is going on. - Two methods to deblur images (generally remove out of focus light from the focal plane)o Laser scanning confocal microscopy uses pinholes to deblur Pinholes can focus length  Gives laser to stimulate fluorescent of image causing it to focus on one specific point when it passes through keyhole When light comes back out, it passes through detector and another pinhole that focuses it as wello Digital deconvolution uses computational methods to deblur Gedownload door Mark Salib


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