bio lab

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created 5 years ago by Giacinta_Yao
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updated 5 years ago by Giacinta_Yao
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1

three lenses for light microscope

condenser, objective, ocular

2

calculate magnification

length of an image/length of the object

3

total magnification

Mobjectives*Mocular

4

reticle ruler

4x: 0.1mm=4ru

10x: 0.1mm=10ru

40x: 0.1mm=40ru

5

resolution

the closest distance that two distinct paralleled edges in the object can be separated

6

contrast

use stain

7

resolution (d)

0.61wavelength/numerical aperture

8

brightfield

produce bright background field of view

9

methylene blue

highlight cell nuclei

10

darkfield

a central dark stop to achieve contrast; shows the components of the cell that have the highest refractive index as bright components against a dark background

11

phase contrast

differences in refractive indices among different components of the specimen produce contrast in the image

12

fluorescence microscope

mole. fluorophores as stains

fluorescence arises because the electrons in the fluorophore mole. can enter several discrete energy levels states. the electrons exist at a low energy. to enter one of the higher energy states requires the absorption of a photon of light. this emitted photon is of a lower energy than the absorbed photon since some energy has already been lost as heat.

fluorophores bleach slowly

13

fluorophores used

DAPI: chromosome, blue, under UV

FITC: tubulin, green, under blue

Rhodamine: actin, red, under green

14

immunofluorescence

the fluorophore is coupled to an antibody: proteins that are produced by the immune sys. to identify and neutralize pathogens.

15

direct immunofluorescence& indirect & primary& secondary

...

16

enzyme

constructed of one or more polypeptide chains held together by covalent bonds

catalyze rxn by a factor of 103/106 by lowering the activation energy

17

active site

fits a specific chemical substrate

form enzyme-substrate complex or transition state

18

cofactors

inorganic, non-polypeptide portions essential to enzymatic act.

19

coenzyme

small organic mole. that baically transport chemical functional grps from one one enzyme-catalyzed rxn to another

20

tyrosinase

a copper-containing enzyme that is involved in melanin synthesis which gives skin its color

21

environmental conditions and enzymatic act.

substrate concentration

T & pH

22

denaturation

the loss of act. while the enzyme remains physically intact

23

spectrophotometer

measure the rate of rxn

24

renaturation

...

25

tyrosinase and DOPA

tyrosine +E +O2-->DOPA+E+O2-->dopaquinone --> dopachrome

26

plasmid

contain the genes to allow for bacterial sex or genes that provide resistance to an antibiotic (not essential)

27

vector

inserted gene

28

pGLO plasmid

insert it into bacteria

29

naturally competent & chemically competent

only a tiny percentage of competent cells will take up the foreign DNA and become transformed

30

transformants

affected cells

31

nontransformants

unaffected bacteria

32

colony

...

33

plate containing ampicillin

only a transformed bacterium can survive and divide

exponential growth from this one cell gives rise to a small circular lump of billions upon billions of cells referred to as a colony

34

pGLO plasmid

engineered plasmid

1) ampicillin resistance gene (bla)

2) GENE THAT CODES FOR THE GREEN fluorescent protein (GFP)

will grow on growth-media-containing ampicillin

35

arabinose operon

only when arabinose (a sugar) is present in the growth media is the GFP gene transcribed. GFP expressed-->transformed bacterial cells will fluoresce under UV light

36

restriction site

the cut site is referred to as the recognition site

37

restriction enzyme

produced by bacteria that cut DNA ole.

38

digested

foreign DNA entering the bacterial cell is digest/hydrolyzed immediately once the restriction site is detected by the restriction enzyme

39

EcoRI

one restriction enzyme extensively used in genetic engineering, the first restrictio nenzyme isolated from E.coli

40

taq polymerase

replicate DNA in vitro

heat-resistant DNA polymerase used to produce virtually unlimited copies of a specific DNA seq in a test tube---PCR

41

PCR

DNA polymerization

1. segment of DNA to be copied

2. the enzyme taq

3. dNTPs, monomers of DNA

4. Primer

a proper buffer solution with a appropriate pH and MG+ cofactor and other salt concentrations is required to provide a suitable chemical environment for optimum act. and stability of the DNA poly

42

PCR process

1. denaturation: 94, separate the two strands

2. annealing 54: hydrogen bonds are formed between bp, between the shot single stranded DNA primers and the complementary regions on the long single stranded DNA template

3. extension 72: near the ideal working T for the enzyme; polymerase adds the complementary bp from the substrate pool of dNTP

43

primer dimer

cross-hybridization w/ untended regions of the template-->amplificatio of those

44

D1S80 LOCUS

short tandem repeat on human chromosome 1

100-300bp

45

plant diversity

1. bryophytes

2. seedless vascular pt

3. gymnosperm

4. angiosperm

46

angiosperm (eudicot/monocot)

eudicot: two cotyledons

leaves usually netlike

4-5 pedals

stem in a ring

root in a central cylinder

47

monocot

one cotyledon

veins usually parallel

usually in multiples of three

stem scattered

root in a ring

48

leaves

both sides of the leaf are enclosed by a layer of epidermal cells coated with a water-proof outer layer called cuticle

upper and lower epidermis (stoma, more lower, CO2 INTO, WATER AND O2 OUT)

palisade mesophyll (chloroplast)

spongy mesophyll

49

ROOT

zone of cell differentiation

elongation

division

root cap

apical meristem