Lecture Summaries (lectures 2,3,4)

what is a cell defined as

structure that carries hereditary information (DNA)

gathers resources and can replicate itself

what is a cell bound by

plasma membrane

who are compound microscopes often attributed to

Janssens

describe the findings of Robert Hooke

observed cork under a microscope and named the empty chambers “cells” after monastery rooms

who was considered a gentlemen scientist

Leeuwenhoek

describe the findings of Leeuwenhoek

using his single lens microscopes with excellent resolution his first observed living bacteria

what did Leeuwenhoek call the living bacteria he saw

animalcules

who proposed the cell theory

Schleiden and Schwann

what were the 3 rules in the cell theory

1. all life is made of 1 or more cells

2. cell is the basic unit of structure

3. all cells arise from pre-existing cells

who added the 3rd rule to the cell theory

Virchow

what idea was Virchow countering

the idea of spontaneous generation

who experimentally disproved spontaneous generation

Louis Pasteur

what are some exceptions/nuances to the cell theory

1. artificial cells

2. viruses (rely on host machinery to replicate)

what are the two methods of creating artificial cells

bottom up (synthesizing the basic building blocks)

top down (remove DNA from existing cell and inserting synthetic genome)

where do all cells store hereditary information

in DNA

how do all cells replicate DNA

semi-conservative replication

what does DNA carry instructions for building

proteins

what are protein catalysts called

enzymes

most reactions in cells do not require enzymes

FALSE

cells translate RNA into protein different ways

FALSE

what is a gene

DNA segment encoding a protein

how do cells obtain the necessary free energy

light or chemical fuels

building blocks of cells are broadly similar across organisms

TRUE

genome size doesn’t vary across living organisms

FALSE

what three fields did cell biology arise from

cytology, biochemistry, genetics

the human eye resolved down to about ____ micrometers

100-200

what is the average size of eukaryotic cells

10-100 micrometers

how big are prokaryotic cells

1-10 micrometers

light microscopes give roughly a ____ fold increase in resolution over the naked eye

1000

electron microscopes give a roughly ____ fold increase in resolution over the naked eye

1 000 000

__ microscopes enable the visualization of individual molecules

electron

electron microscopes resolve down to about _____

200 picometers

light microscopes resolve down to about ____

200 nanometers

approximately how many genes can a minimal bacteria genome require

under 300

magnification alone guarantees more visible detail

FALSE

resolution is the property of that _____

actually determines how much detail can be seen

resolution

the minimum distance between two objects at which they can still be told apart as two separate objects

what is the relationship between object distance and resolution

shorter distance = higher/better resolution

longer distance = lower/worse resolution

what is the resolution equation

resolution = 0.61 x wavelength of light / numerical aperture

the best resolution needs the _____ possible wavelength and the _____ possible NA

shortest, largest

how does oil immersion influence resolution

it increases the refractive index which increase the NA

what light limits light microscopy resolution

visible light (390 nm)

what gives higher resolution

electron beams

what are the different types of light microscopy

1. brightfield

2. phase contrast

3. differential interference contrast (Nomarski)

how does a brightfield light microscope work

light is blocked and absorbed as it passes through the specimen which creates shadows and contrast

what are often needed to see details using a brightfield microscope

stains (cells are mostly translucent)

what are advantagees of using a brightfield microscope

1. inexpensive

2. easy to use

3. can image living samples

how does phase contrast light microscopy work

compares the phase of light that passed through the specimen to light that did not which produces a high contrast image

phase contrast light microscopy produces a high contrast image without stains

TRUE

what are advantages and disadvantages of the phase contrast microscope

advantages

• more detail than brightfield

• works on living cells

disadvantages

• more expensive equipment

• more training

how does the Normarski light microscope work

compares the polarization (rotation) of light that passed through the specimen vs the light that did not

advantages and disadvantages of Normarski

advantages

• produces highly detailed pseudo 3d images

disadvantages

• more complex and costly

• cannot use living cells

how does a fluorescence microscope work

fluorophores absorb light at shorter wavelength (excitation wavelength) and emit it at at longer wavelength (emission wavelength)

compare the excitation and emission wavelengths in a fluorescence microscope

excitation - shorter

emission - longer

how does a dichroic mirror work in fluorescence microscopes

it reflects wavelengths below a cutoff (exciting the sample) and passes wavelengths above the cutoff (emitted light) through to the eyepiece

wavelengths below the dichroic mirror cutoff are known as ____ and wavelengths above the cutoff are known as _____

excited, emitted

where must the dichroic mirror cutoff sit in relation to the excitation and emission wavelengths

between them

fluorescence lets you detect and resolve objects smaller than the resolution limit

FALSE

can you combine multiple dichroic mirrors

YES

what kind of experiment uses multiple dichroic mirrors

co-localization experiment

what are advantages and disadvantages of fluorescence microscopes

advantages

• detects sub-resolution structures

• works with living cells

disadvantages

• expensive equipment and stains

• more user expertise (experience)

what can TEM be essentially seen as

brightfield microscope but using electrons instead of light

what must be done to a sample when using a TEM

must be stained with electron-dense heavy metals

why must samples be stained with heavy metals when using TEMs

so structures can block and reflect the beam to create contrast

what does TEM require

1. vacuum

2. very small samples

3. extensive preparation

4. expensive equipment

advantages and disadvantages of TEM

advantage

• yields very high resolution images but low depth of field

disadvantage

• cannot be used on living cells

• expensive

• requires vacuum

electron microscopy uses ___

magnetic lenses

how does SEM work

electrons are directed at the sample surface and scattered electrons are detected to the side

what kind of image does SEM produce

a high depth of field and 3D looking surface images (NOT 3D)

why are cells so small

• smaller cells have higher surface area to volume ratio = transport of nutrients/waste easier

• larger cells take longer for molecules to diffuse across the cell

• larger cells need more proteins/structural material than DNA can efficiently supply

what are the three domains

bacteria

archaea

eukarya

how are the three domains evolutionary distinct

via a common ancestor (LUCA)

what two domains are grouped together as prokaryotes

bacteria and arcaea

____ Lack a true nucleus; _____ have a true nucleus

prokaryotes, eukaryotes

archaea are metabolically similar to ____ and handle genetic information more like ______

bacteria, eukaryotes

__ lack membrane bound organelles

prokaryotes

is the inside of prokaryotes organized

yes

____ are compartmentalized

eukaryotes

what defines the boundary of the cell

plasma membrane

why do cells compartmentalize

• prevents interference between opposing cellular processes occurring at the same time

• increases efficiency of resource use

what is the trade off in cell compartmentalization

compartmentalized systems are more complex = require membrane maintenance and transport mechanisms = requires bigger genomes and lower replication

what is the first step in studying organelles with centrifugation

cells first must be homogenized (membranes ruptured) to release contents

compare differential centrifugation and density gradient centrifugation

differential

• particles settle at different rates depending on size, density, and shape where successive spins at increasing speed and time precipitate out larger particles then smaller ones

density

• sample is layered on top of a gradient of increasing density (down the test tube) and particles migrate until they reach the surrounding liquid that matches their own density

what centrifugation method yields a high enriched but not pure sample

differential centrifugation

how did the nucleus and ER originate

likely arose from invagination (moving inward) of the plasma membrane around the DNA which created a double membrane around the nucleus

how did chloroplasts and mitochondria originate

originated when a prokaryote was engulfed by an eukaryotic cell

how many membranes do mitochondria have? why?

2 - one from the engulfed prokaryote and one derived from the host’s membrane

endosymbiotic theory is essentially based on

an ancient ‘accident’ that became a stable symbiotic relationship