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microbiology > chemistry + microscopy 9/14 > Flashcards

chemistry + microscopy 9/14 Flashcards

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Biology 101 flashcards
1
Q

what are differences between DNA and RNA

A

§DNA

Double-stranded in cells, complementary strands

Strands held together by hydrogen bonds

§RNA

Typically single-stranded

Demonstrates secondary structure (folding back upon itself)

Four classes: mRNAs, tRNAs, rRNAs, and small RNAs

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2
Q

what are the two major types of proteins

A

§Enzymes - catalytic proteins; catalysts for chemical reactions

§Structural Proteins

Integral parts of cellular structures (such as eukaryotic chromosomes)

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3
Q

amino acids (monomers) characteristics

A

§Most consist of carbon, hydrogen, oxygen, and nitrogen; 2 of 22 contain sulfur, 1 contains selenium

§All contain two important functional groups

§Carboxylic acid group (-COOH)

§Amino group (-NH2)

§Adjacent amino acid monomers held together by covalent bonds called peptide bond

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4
Q

what are optical isomers

A

§optical isomers = enantiomers (sterioisomers or mirror image isomer); have same chemical properties but often have different physical properties

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5
Q

what are racemases

A

§enzymes capable of interconverting specific enantiomers

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6
Q

what is a polypeptide

A
  • structural term meaning a series of amino acids (10s,100s,1000s or 10,000s amino acids long) joined to each other by peptide bonds
  • Each polypeptide has an amino end and a carboxyl end
  • A given polypeptide could be a whole protein or only a subunit of a larger protein
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7
Q

what is a protein

A

a functional unit consisting of one or more polypeptides having one or more functions.

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8
Q

every protein has at least one

A

polypeptide

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9
Q

what is primary structure

A

§linear array (sequence) of amino acids in a polypeptide

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10
Q

what is secondary structure

A

§localized folds or twists in parts of polypeptide that form a more stable structure; held together by hydrogen bonding between amino group Hydrogen and carbonyl Oxygen

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11
Q

what is teriary structure

A
  • overall, 3 dimensional shape of a polypeptide
  • Forms exposed regions or grooves in the molecule (i.e., active site) that are important for binding to other molecules
  • Held together by 1) hydrogen bones, 2) electrostatic interactions, 3) hyrophobic interactions with water and 4) Disulfide (strong) bonds: covalent bonds between -SH groups from two different amino acids
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12
Q

what is quaternary structure and a subunit

A
  • overall arrangement of polypeptides in a protein; only found in proteins composed of two or more polypeptides
  • each polypeptide in the protein, held together by either/both covalent and noncovalent linkages
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13
Q

what is the difference between gentle and harsh denaturation methods

A

gentle methods don’t break covalent bonds so the protein can be put back together if necessary while harsh methods break the covalent bonds

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14
Q

what is denaturation

A

unfolding of polypeptide chains causing loss of biological function

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15
Q

what is denaturation caused by

A

breaking of specific bonds from extreme pH, high temperatures, certain chemicals

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16
Q

what do microscopes do and what do they vary in

A
  • produce magnified images of objects
  • vary in

–1) Illumination source: light; electrons, other

–2) Focusing method: glass lenses; magnets, other

–3) Specimen preparation

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17
Q

what are light microscopes and what are the two types

A
  • use visible light to illuminate
  • use glass lenses to focus
  • have light pass through/around specimen
  • Simple = 1 focusing lens
  • Compound = 2 focusing lenses form the image
  • objective lens and ocular lens
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18
Q

what is total magnification

A

product of magnification of two sets of lenses (–Objective magnification X ocular magnification; maximum is about 2,000)

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19
Q

what is resolution

A

•the ability to distinguish two close, adjacent objects as separate and distinct

  • determined by wavelength of light used and lenses used
  • shorter wavelength = greater resolution
  • Limit of resolution for best light microscopes is about 0.2 mm
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20
Q

what are the difference varieties of light microscopes

A

–Bright-field microscope

–Dark-field microscope

–Phase-contrast microscope

–Fluorescence microscope

–Confocal microscope

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21
Q

what is a bright field microscope

A
  • common
  • Specimens are visualized because of differences in contrast (density) between specimen and surroundings
  • Bright-field due to specific condenser lens

–Entire field illuminated

  • objective and ocular lenses
  • produces dark image against brighter background
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22
Q

what is working distance

A

distance between the front surface of lens and surface of cover glass or specimen when it is in sharp focus

23
Q

why is oil used

A

If air is replaced with immersion oil, many light rays that did not enter the objective due to reflection and refraction at the surfaces of the objective lens and slide will now do so. This results in an increase in resolution and numerical aperture.

24
Q

what does the bright field microscope usually require

A

•staining objects to achieve sufficient contrast between specimen and surrounding medium

25
Q

what are photos taken by bright field microscopes called

A

photomicrographs

26
Q

what is the dark field microscope

A
  • Uses special condenser lens
  • Produces a bright image of the object against a dark background
27
Q

what is the dark field microscope used to observe

A

used to observe living, unstained preparations - –used to observe internal structures in eukaryotic microorganisms –used to identify bacteria such as Treponema pallidum, the causative agent of syphilis

28
Q

what is the phase contrast microscope

A
  • Uses special condenser lenses
  • Converts slight differences in refractive index and cell density into easily detected variations in light intensity
  • Some light rays from hollow cone of light passing through an unstained cell are retarded and out of phase and dark compared to the bright background
29
Q

what is the phase contrast microscope used to observe

A

•Used to observe living cells

–studying microbial motility

–detecting bacterial structures such as endospores and inclusion bodies that have refractive indices different from that of water

30
Q

what is the fluorescence microscope used for

A
  • Used to visualize specimens that fluoresce after exposure specimen to ultraviolet, violet, or blue light
31
Q

fluorescence microscope characteristics

A
  • Some cells fluoresce naturally (autofluorescence); others after they have been stained with a fluorescent dye
  • Shows a bright image of the object resulting from the fluorescent light emitted by the specimen
  • Has applications in medical microbiology and microbial ecology studies
32
Q

how does fluorescence microscopy work

A

–fluorochrome-labeled probes, such as antibodies, or fluorochrome dyes tag specific cell constituents for identification of unknown pathogens

–localization of specific proteins in cells

33
Q

how does the differential interference microscope work

A
  • Creates image by detecting differences in refractive indices and thickness of different parts of specimen
  • Excellent way to observe living cells

–live, unstained cells appear brightly colored and three-dimensional

–cell walls, endospores, granules, vacuoles, and nuclei are clearly visible

34
Q

how do unstained slide preparations work

A

–Wet mounts – drop of liquid with living organisms; unstained

•Hanging drop method; concave slides

35
Q

how does a stained slide preparation work and what are the types

A

–Vital Stains – dye added to wet mount; stains living organisms; may eventually kill microorganism

–Fixed and stained slides – kills microorganisms

36
Q

how can contrast be improved in light microscopy

A
  • Improving contrast results in a better final image
  • Staining improves contrast between specimen and surrounding

–Dyes or stains are compounds that bind to specific cellular materials

–Examples of common stains are methylene blue, safranin, and crystal violet

37
Q

what is fixation

A

•kills microorganisms, preserves internal and external structures and adheres them to the slide

38
Q

what is an artifact

A

any distortion in the true morphology of the microorganism due to the technique itself

39
Q

what are the different types of fixation

A

–1) Heat fixation – bunsen burner; artifacts;

–2) Chemical fixation – chemicals, more expensive, more time, less artifacts; used with larger, more delicate organisms

•protects fine cellular substructure and morphology

40
Q

what are stains/dyes

A

–organic compounds or salts that have different affinities for specific cellular materials

-most are salts of cation and anion

41
Q

what is a chromophore and the difference between a basic and acidic dye

A

•Chromophore - coloring ion

  • Basic dye – chromophore cation; binds to cells; most dyes
  • Acidic dye – chromophore anion; repelled by cells; stains background
42
Q

what do dyes do

A

–make internal and external structures of cell more visible by increasing contrast with background

43
Q

what is the general idea of a simple stain and the steps

A

•1 dye; few steps; all stained the same

–use can determine size, shape, and arrangement of bacteria

  • steps
    • prepare a smear by spreading culture in thin film over slide and drying in air
    • heat fix and flood slide with stain - rinse and let dry
    • examine with microscope
44
Q

what are differential staining techniques and what do they do

A
  • 2 or more dyes; several steps
  • Divides microorganisms into groups based on their staining properties

–e.g., Gram stain

–e.g., acid-fast stain

•Differential stain used to detect presence or absence of structures

–endospores, flagella, capsules

45
Q

what is gram staining

A
  • Most widely used differential staining procedure
  • Divides bacteria into two groups, Gram-positive and Gram-negative, based on differences in cell wall structure
46
Q

about acid staining

A

•Particularly useful for staining members of the genus Mycobacterium

e. g., Mycobacterium tuberculosis – causes tuberculosis
e. g., Mycobacterium leprae – causes leprosy

–High lipid content in cell walls (mycolic acid) is responsible for their staining characteristics

47
Q

what is endospore staining

A

–heated, double staining technique

–bacterial endospore is one color and vegetative cell is a different color

48
Q

what is a capsule stain

A

•used to visualize capsules surrounding bacteria

–negative stain - capsules may be colorless against a stained background

49
Q

what is flagella staining

A

–mordant applied to increase thickness of flagella

50
Q

what is electron microscopy

A

•Uses electrons instead of light as the illuminating beam

–Illumination – electron beam

–Lenses – magnets

–Image – photograph or computer

–Stains – metals

  • Wavelength of electron beam is much shorter than light, resulting in much higher resolution
  • Allows for study of microbial morphology in great detail
51
Q

about transmission electron microscope

A
  • Beam of electrons pass through specimen
  • Electromagnets function as lenses
  • Electrons scatter when they pass through thin sections of a specimen
  • Transmitted electrons are under vacuum which reduces scatter and are used to produce clear image
  • Denser regions in specimen, scatter more electrons and appear darker
  • Enables visualization of structures at the molecular level
  • Specimen must be very thin (20–60 nm) and be stained with metalic stains
52
Q

how are specimens prepared for electron microscopy

A
  • Analogous to procedures used for light microscopy
  • For transmission electron microscopy, specimens must be cut very thin
  • Specimens are chemically fixed and stained with electron dense materials, such as heavy metals, that differentially scatter electrons
53
Q

what are other methods of electron microscopy prep

A

•Negative stain

–heavy metals do not penetrate the specimen but render dark background

–used for study of viruses, bacterial gas vacuoles

•Shadowing

–coating specimen with a thin film of a heavy metal on only one side

–useful for viral morphology, flagella, DNA

•Freeze-etching

–freeze specimen then fracture along lines of greatest weakness (e.g., membranes)

–allows for 3-D observation of shapes of intracellular structures

–reduces artifacts

54
Q

what is the scanning electron microscope and how does it work

A
  • Uses electrons reflected from the surface of a specimen and collected by a detector to create detailed image
  • Produces a realistic 3-dimensional image of specimen’s surface features
  • Can determine actual in situ location of microorganisms in ecological niches
  • Magnification range of 15´–100,000´

microbiology (11 decks)

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