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BIOL 243 > Lab exam Review > Flashcards

Lab exam Review Flashcards

1
Q

Why do we add diswashing detergent to extract DNA? (4)

A

The detergent lyses the cell membrane and denatures proteins (by breaking protein-protein interactions). Increase membrane permeability by removing lipids, break cell membrane as detergents bind to hydrophobic bilayer and water

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2
Q

Why do we add table salt in DNA extraction? (3)

+Config

A
  • The positively charged sodium ions in table salt form
    favourable electrostatic interactions with the negatively charged DNA, whereas the negatively charged chloride ions form favourable electrostatic interactions with the positively charged histones.
  • This helps neutralize DNA and less hydrophillic so it doesnt disolve in water.
  • The salt also help remove proteins that are bound to the DNA
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3
Q

Why do we add isopropanol?

A

To get DNA out of solution we add isopropanol to decrease the polarity of the water and makes DNA less soluble.

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4
Q

SYBR Gold

A

A flourescent dye that gives off bright yellow-gold
light when bound to DOUBLE STRANDED DNA and exposed to blue light.

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5
Q

Procedure for strawberry DNA extraction LAB 1

10 steps

A
  1. crush a thawed strawberry in ziploc
  2. Add DNA buffer which includes detergent
  3. strain DNA with cheesecloth over falcon tube (make sure no fibre)
  4. While filtering, label 3 microcentrifuge tubes X,Z and add (10,20ul) of water. Tube Y has already been prefilled for you and contains 10 μL of 2M NaOH
  5. In the DNA tube pour ice-cold isopropanol into the tube
  6. Dip a Pasteur pipette so DNA can attach
  7. stir in ethanol 10 times with the pipette to help wash away some of the contaminant proteins, lipids and salts attached to the surface of the DNA.
  8. dip into tube b to dissolve dna
  9. pour allqouit into centrifuge tube
  10. centrifge for pellet
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6
Q

how centrifuge work

A

a machine that uses centrifugal force to separate the contents of a sample based on their density

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7
Q

For Lab 1 what would you expect to see in tube x,y,z?

A

X:water+DNA+Dye so it would florescent the brightest
Y:Dye+NaOH+DNA it would florescent less
z:DYE+water no flourescent as no DNA is present

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8
Q

What does NAOH do to DNA in Lab 1?

+what happpens with single stranded?

A

Sodium hydroxide is a strong base that removes some of the protons in the nitrogenous bases of DNA, thus disrupting the hydrogen bonding needed to stabilize the structure of double-stranded DNA. SInce dye only bind/flourescent to double stranded DNA it would not flourscent with single stranded. Single-stranded/DNA does not bind strongly to the dye because the bases are not stacked tightly in this form
of DNA

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9
Q

What is in the 15% saline nutrient agar?

A

Nutrient agar with 15% NaCl

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10
Q

How are the salinity gradient agar plates made and how do they work? (4)

A

When you pour the first salinity agar, the plate is tilted by a 5mm glass rod so the raised end is low salinity concentration and the low end is thicker with the salinity agar.

You then add the molten nutrient agar with the plate flat on the bench.

The salt from the lower wedge diffuses into the upper non-saline layer, generating a linear gradient of salt concentration across the upper layer.

Bacteria inoculated on the surface are
thus exposed to varying levels of salinity across the Petri dish.

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11
Q

The arrow on the petri dish is pointing to

A

the thinnest depth of agar aka lowest salt concentration

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12
Q

The second layer nutrient agar takes longer to set because

A

of the insulating effect of the first layer

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13
Q

Chi sqaure test 🤢 (3)

What does the chi square determine for our results+null hypoethsis?

A
  • How well observed ratio fit expected ratio
  • Chi sqaure test tells us if the deviations between the observed data and our predicted results are due to chance alone or if other factors are responsible
  • Tells you the probability (expressed in percent) that chance alone has caused the deviation between observed and expected results. If chance has caused the difference between observed and expected results, then the results fail to reject the null hypothesis.
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14
Q

When performing Chi sqaure tests, our null hypothesis is:

A

There is no statistically significant difference in any differences we observe (i.e., that any differences are due to
chance alone

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15
Q

Chi sqaure null hypothesis for coin toss:

A

There will be no difference between the observed number of heads and tails and the expected number of heads and
tails, and that any deviation of the observed results from the expected results is due to chance alone, and not to some other cause.

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16
Q

The greater the discrepancy between the observed and expected values, the…..

x2

A

larger the value of χ^2

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17
Q

Chi sqaure formula

A
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18
Q

If x^2 caculated is greated then the X^2 value in the table then

A

the difference between the observed and expected resylts us not due to chance alone and is statistcally signifigant

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19
Q

If x^2 caculated is smaller then the X^2 value in the table then

A

the difference between obersved and expected results is likely due to chance alone and the difference is not statistically signifigant

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20
Q

A small P value reference indicates

A

That you are less likely to reject
your null hypothesis when it is in fact true (only a 1 in 100 chance) but it also means that you are also less likely to reject the null hypothesis when it is wrong.

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21
Q

Degree of freedom

A

One less than the number of categories of possible outcomes (n − 1). For our coin toss example, we have two categories of
possible outcomes (i.e., heads or tails), so there is (2 − 1) = 1 degree of freedom.

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22
Q

If you had calculated a χ2 value of 0.37 with 1 degree of freedom, the probability would lie
between 50% (0.455) and 80% (0.0642). This tells you that

A

deviation of 0.37 from expected values will occur by chance alone in 50%–80% of trials. You would be very confident that there is no significant difference between your observed and expected results and you would fail to reject the null hypothesis.

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23
Q

Statistically significant means a result is

A

unlikely due to chance (i.e., sampling error)

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24
Q

How does salinity influence the growth and survivial of organisms?

A

through the effect of salts on osmotic balance, as well as direct effects on proteins. Only a few organisms can survive in extremely high saline environments.

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25
Q

In our bacteria evolution lab what are we comparing?

A

Comparing bacterial cell cultures that
have been exposed to a condition that can increase the rate of mutation with unexposed cells, to determine if increasing the rate of mutation has an effect on the adaptive response to increasing salinity.

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26
Q

Inoculating

A

To introduce something such as bacteria, a virus, or a fungus into an animal or plant as part of an experiment or to encourage it to grow there

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27
Q

How did we prepare our S1 and C1 plates?

A

We used bacteria in nutrient broth and placed 6 droplets

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28
Q

What is the sat concentration throught left to right hand side?

Arrow point to right hand side

A

The salinity gradient in the nutrient agar layer across the Petri dish ranging from 15% NaCl at the left-hand edge of the Petri dish to 0% on the right-hand side.

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29
Q

How do you find the approximate saline concentration in the region of the plate where the colony is growing?

A

You will need to measure the diameter of the Petri
dish. If you divide the maximum saline concentration by the diameter of the plate, you will know the change in saline concentration per millimeter as you move across the plate.

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30
Q

Control plate

A

No saline

Prepared for us**

31
Q

How did we prepare our S2 and C2 plates?

A

We streaked our colony S1 and C1 plates (with the six droplets) from the colony that is growing closest to the left-hand edge (i.e., the highest saline concentration) of your S1 plate. Then we transfered to our S2 plat by streaking where the agar has the highest saline concentration and will end where the agar has the lowest saline concentration.

32
Q

How did we prepare our S3 and C3 plates?

A

We pick up a sample from the colony that is growing closest to the left-hand edge (i.e., the highest saline concentration) of your S2 plate. And then

33
Q

Sample mean

What is this an estimate of?

A

The mean for the sample of organisms that you used in your experiment.
This sample mean is an estimate of the population mean, the value that you would obtain if you could have included every organism of the species on the planet.

34
Q

What do T test do?

A

A statistical test that allows us to determine whether two means are significantly different

It is often used in hypothesis testing to determine whether a process or treatment actually has an effect on the population of interest, or whether two groups are different from one another.

35
Q

The larger the T test value….

A

the more likely it is that the difference between the two means is statistically significant.

36
Q

Why do the isopronanol have to be ice cold in strawberry lab?

A

Because enzyme DNAses, like other enzymes are temperature sensitive and therefore DNAses are not active when the temperature is too low. Since these enzymes are involved in DNA lysis & degradation, reduction in this enzyme’s activity will yield more amount of pure DNA.

37
Q

For lab 1 what are we testing and how?

Key words: SYRB, SSDNA,

A

DNA does not absorb visible light very well, so it is basically colourless to the naked eye. However, SYBR Gold bind strongly to the negatively charged phosphates and hydrophobic bases of DNA , it only gives off fluorescent light when bound to double-stranded DNA. The dye interacts with both the negatively charged backbone and the closely stacked bases found in double-stranded DNA. In contrast, single-stranded DNA does not bind strongly to the dye because the bases are not stacked tightly in this form of DNA. We will confirm this property by visually estimating the amount of fluorescent light given off when the dye is added to either water, DNA, or DNA denatured by the addition of a small amount of sodium hydroxide.

38
Q

Falcon tubes

A
39
Q

Transformation

A

a process in which a bacterial cell takes up DNA from its
environment.

40
Q

Origin of replication (ori):

A

Bacterial DNA sequences that are required for replicating the
plasmid prior to cell division

41
Q

Gene of interest (GFP):

A

DNA sequence from the jellyfish A. victoria that provides the protein-coding sequence of GFP

42
Q

Promoter

A

DNA sequence from a bacterial gene promoter that allows for the production of messenger RNA (i.e., transcription) from the GFP gene;

43
Q

Inducer (araC)

A

DNA sequence from a bacterial gene regulatory operator that allows transcription of the GFP gene only when the sugar arabinose is present;

44
Q

Selectable marker (ampR):

+What it is+what it does +How it does it

A

DNA sequence from a bacterial source that allows the transformed bacterium to make the enzyme β-lactamase, an enzyme catalyzes the breakdown of ampicillin

This DNA sequence confers ampicillin resistance to transformed bacteria

45
Q

Ampicilin

A

an antibiotic that normally prevents E. coli from growing.

46
Q

Five different DNA sequence elements in the pGLO plasmid, which are carefully arranged to allow for the synthesis of GFP in E. coli

A
  1. Origin of replication (ori):
  2. Gene of interest (GFP):
  3. Promoter
  4. Inducer (araC)
  5. Ampr
47
Q

Whats in the +PGLO tube? (4)

A
  1. plasmid DNA
  2. Transformation solution (CaCl2)
  3. E coli cells
  4. LB broth
48
Q

The plasmid that you are treating your E coli cell contains the gene for 2 proteins:

A
  1. Amp resistance
  2. GFP gene
49
Q

The GFP gene is under the control of a—— that will only be turned on in the prescence of ——

A

promotor
arabinose sugar

50
Q

The E coli with transformed plasmid can

Not abt the GFP gene

A

grow on LB agar that contains amplicilin

51
Q

Role of Cacl2 in the transformation solution:

A

Ca2+ role is to neutralize natural negative charges beared both by DNA and cell surface(mainly LPS) to avoid electrostatic repulsion between the two.
to counteract the electrostatic repulsion between the plasmid DNA and bacterial cellular membrane

52
Q

Yeast lab one describe to me the procedures

A
  1. spread yeast (either wildtype or light sensitive) solution on the agar plates
  2. Obtain four cardboard templates from the back bench, one for each Petri dish, and four pieces of Saran wrap just big enough to cover the template.
  3. Apply sunscree 1, 2 and foil to the three plate and none for fourth
  4. Expose the treatments to UV light
53
Q

Results of the UV yeast lab (5)

Overall trend, under full radiation, colony amount, why?

A
  • there is variation between the two individual strain as SEM did not overlap between wildtype and light sensitive bars
  • The wildtype showed greated genectic stability and higher success of DNA repair under full UV radiation
  • Largest colony was found for both strain on the foil treatment
  • Lowest SPF had the lowest cell colonies
  • UV radiation affect DNA replication causes disortion to structure and lesions
54
Q

If the mutated codon encodes for the same amino acid, this is called….

A

synonymous mutation/silent mutation

55
Q

If the mutated codon encodes for a different amino acid, this is called

A

non-synonymous mutation/missense
mutation

56
Q

If the mutated codon encodes a stop codon, write “stop”, this is called

A

nonsense mutation

57
Q

Why did we uncubate the microcentrifuge tubes after heat shocking it? (plasmid+Ecoli already added)

A

So that there is time for AmpR expression to produce beta-lactomase
Following heat shock or electroporation, transformed cells are cultured in antibiotic-free liquid medium for a short period to allow expression of antibiotic resistance gene(s) from the acquired plasmid to begin

58
Q

How does sugar arabinose activate GFP?

A
59
Q

How was the control onion root prepared?

A

The Control set of root tips were incubated in water for 24 hours at 4°C in the dark.

60
Q

How were the treatment onion root prepared?

A

incubated in one of three different beverages, each containing
different amounts of caffeine, for 24 hours at 4°C in the dark.

61
Q

What does HCL do while preparing the onion root top?

What does it do, how (does two thing to contribute to the one thing)

A

The HCl helps break apart the tissue so the cells can be seen better.) Break up the cellulose cell wall allowing stain to permeate the tissue and makes it easier to squash the tissue on a microscope slide

62
Q

How were the onion root cell lab prepared?

A
  1. Add the root to 0.5ml of 1M HCL and incubate
  2. take out root and wash HCL away by dripping water ontop
  3. Add Toluidine Blue (dye)
  4. Add water to wash away excess dye
  5. Add 45% acetic acid
  6. Add water droplet
63
Q

male heterozygous for both gene A and gene
B would be written as:

A

A/a ; B/b ; X/Y

64
Q

metacentric chromosome

A

has arms on either side of the centromere of equal length.

65
Q

acrocentric chromosome

A

as one arm longer on one side of the centromere than the other

66
Q

acrocentric chromosome

A

as one arm longer on one side of the centromere than the other

67
Q

Albino

A

recessive mutation and individuals that are
homozygous for the a allele (a/a) do not have any skin
pigmentation

68
Q

chi sqaure and t sqaure diff ur own worlds lmao

A

Chi sqaure: does my result deviate from whats suppose to happen?

T-test: do the mean treatments i put on have an effect on this other mean one?

69
Q

In text citation for John Smith at 1990

A

The high infant mortality rate in the U.S. may be attributed in part to the high cost of medical insurance (Smith 1996). Smith (1996) found that economic barriers to adequate prenatal …

70
Q

In text citation for no author name but 2006 pulished

A

Some people have found vitamin E helpful for this problem (Ways to overcome … 2006).

71
Q

In text citation from secondary source (Wright is using a source from Frost and Krahn in a 2000 paper on page 8 in his 1999 paper)

A

Wright (1999) argues that drug companies “hold governments hostage” when they refuse to justify the cost of life-saving but highly expensive medications (as cited by Frost and Krahn 2000 p. 8).

72
Q

For in text citations there are no —- except for

A

”.” except for et al.

73
Q

Literatire cited

A

last name-space-first inital-“comma repeat for more authors”-Period-publication year-period-journal title-period-“(volume number in bracket)”-colon-page range

74
Q

Degree of freedom for t test

A

sum of all individual sample size- number of sample sizes

BIOL 243 (22 decks)

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